Elsevier

Brain Research

Volume 890, Issue 1, 26 January 2001, Pages 118-129
Brain Research

Research report
ATP-sensitive potassium channels (KATP) in retina: a key role for delayed ischemic tolerance

https://doi.org/10.1016/S0006-8993(00)03152-8Get rights and content

Abstract

The objectives of the present study were to determine the localization of KATP channels in normal retina and to evaluate their potential roles in ischemic preconditioning (IPC) in a rat model of ischemia induced by increased intraocular pressure (IOP). Brown Norway rats were subjected to sublethal 3-, lethal 20- and 40-min ischemia and the functional recovery was evaluated using electroretinography. The time interval between ischemic insults ranged from 1 to 72 h. The effects of KATP channel blockade on IPC protection were studied by treatment with 0.01% glipizide. IPC was mimicked by injection of KATP channel openers of 0.01% (−)cromakalim or 0.01% P1060 72 h before 20-min ischemia. Co-expression of KATP channel subunits Kir6.2/SUR1 was observed in the retinal pigment epithelium, inner segments of photoreceptors, outer plexiform and ganglion cell layers and at the border of the inner nuclear layer. In contrast to a 20- or 40-min ischemia, a 3-min ischemia induced no alteration of the electroretinogram (ERG) and constituted the preconditioning stimulus. An ischemic challenge of 40 min in preconditioned rats induced impairment of retinal function. However, animals preconditioned 24, 48 and 72 h before 20-min ischemia had a significant improvement of the ERG. (−)Cromakalim and P1060 mimicked the effect of IPC. Glipizide significantly suppressed the protective effects of preconditioning. In conclusion, activation of KATP channels plays an important role in the mechanism of preconditioning by enhancing the resistance of the retina against a severe ischemic insult.

Introduction

Ischemic preconditioning (IPC) is a powerful protective mechanism against ischemic injury that has been shown to occur in a variety of organ systems, including heart, brain, spinal cord, liver, lung, skeletal muscle and recently, retina [9], [21], [29], [35], [42]. Ischemic preconditioning has both immediate and delayed protective effects, the importance of which varies between species and organ systems. While the exact mechanisms of both protective components are yet to be clearly defined, ischemic preconditioning is a multifactorial process requiring the interaction of numerous signals, second messengers and effector mechanisms.

ATP-sensitive potassium channels (KATP channels) couple cell metabolism to membrane potential in many organs [3]. The pancreatic-cell KATP channels play a critical role in the regulation of glucose-stimulated insulin secretion [7], [12]. KATP channels shorten the action potential in hypoxic cardiac muscle and provide one pathway for vasodilatation in vascular smooth muscle [13], [14], [16], [39]. They are also characterized by their inhibition by antidiabetic sulfonylurea drugs such as glibenclamide, glipizide, tolbutamide [16], [45] and their activation by K+ channel openers (KCOs) such as (−)cromakalim, pinacidil, diazoxide [13], [14], [39]. Although the role of KATP channels in the mechanism of myocardial IPC is still controversial depending on the species (for review see Ref. [36]), a considerable body of evidence has implicated KATP channels in cerebral protection. Opening of KATP channels with KCOs protect neurons against the deleterious effects of excitotoxicity and ischemia [1], [22]. Recently, KATP channels have also been shown to be involved in epileptic and ischemic brain tolerance [23], [38]. KATP channels are made of a complex of two structurally distinct subunits, an inwardly rectifying potassium channel subunit (Kir6.x) forming the channel pore [26] and a regulatory subunit, the sulphonylurea receptor (SUR) belonging to the ATP-binding cassette superfamily [2], [25], [27]. A total of five isoforms of SURs have been cloned, SUR1 and four spliced variants of SUR2, SUR2A–D. Probably because the expression and distribution of KATP channels in retinal cells are not yet known, their physiological function in retina is still not well understood.

The aims of the present work were to determine the localization of KATP channels in normal retina and evaluate their potential roles in ischemic preconditioning in a rat model of ischemia induced by increased intraocular pressure (IOP).

Section snippets

Animals

Adult male Brown Norway rats weighing 250 g (Charles River Breeding Laboratory, St Aubain les Elbeuf, France) were housed in clear plastic cages in a room with controlled temperature (22±1°C) and a fixed 50-lx fluorescent (Philips, France) lighting schedule (lights on from 08:00 to 20:00 h). Food and tap water were provided ad libitum. Rats were acclimated for 1 week before experiments. Before electroretinogram (ERG) recordings, the animals were kept in total darkness for a minimum of 16 h.

Distribution of KATP channels in retina

The sensitive reverse transcription-PCR (Fig. 2A) shows that SUR1 and KIR6.2 were detected in retina, whereas the expression of SUR2 was not detectable. In situ hybridization analysis confirms this result. As shown in Fig. 2C, Kir6.2 is highly expressed in retinal pigment epithelium, inner segments of photoreceptors, outer plexiform, inner nuclear and ganglion cell layers, and probably Müller cells. The SUR1 mRNA hybridization is predominantly confined to retinal pigment epithelium, inner

Discussion

Four major findings emerge from the present experiments: (i) the sulphonylurea receptor SUR1 and the inward-rectifier K+ channel subunit Kir6.2 are highly expressed in the control retina; (ii) as previously described in a model of retinal ischemia induced by optic nerve bundle ligation [42], ischemic preconditioning induces impressive retinal tolerance to ischemia in a model of increased intraocular pressure; (iii) the mechanism of the retinal tolerance involves the early activation of KATP

Acknowledgements

This work was supported by the Centre National de la Recherche Scientifique (CNRS), the Association pour la Recherche sur le Cancer (ARC), the Conseil Regional (PACA), the Association Française contre la Myopathie (AFM) and the Fondation de l’Avenir. We thank C. Widmann, N. Vaillant, G. Jarretou, F. Aguila and V. Lopez for technical assistance.

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