Sustained elevation of extracellular adenosine and activation of A₁ receptors underlie the post‐ischaemic inhibition of neuronal function in rat hippocampus in vitro

Abstract

Adenosine is released from the compromised brain and exerts a predominately neuroprotective influence. However, the timecourse of adenosine release and its relationship to synaptic activity during metabolic stress is not fully understood. Here, we describe experiments using an enzyme-based adenosine sensor to show that adenosine potently (IC 50 1 lM) inhibits excitatory synaptic transmission in area CA1 during oxygen/ glucose deprivation ('ischaemia'), and that the prolonged postischaemic presence of extracellular adenosine sustains the depression of the field excitatory postsynaptic potential (fEPSP). N-methyl-D-aspartate (NMDA) receptor antagonism promotes post-ischaemic recovery of the fEPSP, in parallel with reduced release of adenosine. Paradoxically, however, after ischaemia the fEPSP recovers in the face of concentrations of adenosine capable of fully eliminating synaptic transmission during ischaemia. This hysteresis is not prevented by NMDA receptor antagonism, is observed during repeated ischaemia when adenosine release is reduced, and does not reflect desensitization of adenosine A 1 receptors. We conclude that adenosine exerts powerful inhibitory actions on excitatory synaptic transmission both during, and for some considerable time after, ischaemia. Therapeutic strategies designed to exploit both the continued presence of adenosine and activity of A 1 receptors could provide benefits in individuals who have suffered acute injury to the CNS. Keywords: adenosine, epilepsy, hypoxia, ischaemia, neuroprotection, stroke. Adenosine is released during metabolic or traumatic insults to the mammalian, including human, brain and is widely regarded as exerting a neuroprotective influence. Adenosine acts via four G-protein-coupled receptors (A 1 , A 2A , A 2B and A 3 ), of which the A 1 receptor is expressed at the highest level in brain and provides both an important inhibitory and neuroprotective influence (Fredholm et al. 2005a). Various studies have shown that A 1 receptor agonists protect neurones from injury whilst A 1 antagonists generally exacerbate injury (Rudolphi et al. 1992). Similarly, increasing extracellular adenosine by preventing adenosine uptake or breakdown is also neuroprotective, whilst promoting adenosine metabolism worsens neuronal injury Stone 2002). These observations have led to the concept that adenosine, via inhibition of glutamate release and neuronal activity, is an endogenous neuroprotective compound. Conversely, similar pharmacological approaches have shown that the acute administration of A 2A and A 3 receptor agonists worsen, and antagonists improve, the outcome after cerebral ischaemia. One explanation for the contrasting actions of A 2A and A 3 receptors is that their activation limits the beneficial effects of the A 1 receptor, perhaps by inducing adenosine A 1 receptor desensitization (Pearson et al. 2003;Fredholm et al. 2005a).Considerable information regarding the consequences of adenosine receptor activation has been amassed using a

Role of adenosine A3 receptors on CA1 hippocampal neurotransmission during oxygen–glucose deprivation episodes of different duration

“…Alterations in AD characteristics caused by A 3 antagonists may be attributable to their actions on glutamate-mediated cellular responses. The time window of A 3 receptor-mediated effects found in the present work and reported by Pugliese et al [14] overlaps with the delay that can be obtained by treating the slices with glutamate receptor antagonists [25,40,41]. NMDA receptors are essential to AD initiation and propagation [26].…”

“…Therefore, during a prolonged severe OGD insult, the roles of A 1 and A 3 receptors drastically diverge. On the whole, the data support the wellestablished neuroprotective role of A 1 receptors [9,12,41] and confirm an opposite, deleterious, role of A 3 receptors during prolonged ischemia [14].…”

“…During hypoxic/ischemic conditions, adenosine is released from hippocampal slices, reaching concentrations up to 30 μM after 5-min OGD [10,41]. However, the efficacy of A 3 antagonists in blocking the depression induced by OGD of different durations indicates that endogenous adenosine concentrations reached in the extracellular milieu alone are not sufficient to desensitize A 3 receptors.…”

The adenosine kinase hypothesis of epileptogenesis

“…SE, ischemia, brain injury) ADK is rapidly downregulated leading to elevated levels of adenosine, thus enhancing the protective functions of the adenosine system via increased activation of A 1 Rs. Thus, the induction of ischemia in hippocampal slices led to the rapid increase in extracellular adenosine within 10 min to levels >20 µM above baseline (Pearson et al, 2006). This rapid rise of adenosine was demonstrated to be due to the release of adenosine as such and was not dependent on extracellular cleavage of ATP (Frenguelli et al, 2007).…”

“…While A 1 and A 2A receptors have important functions under physiological conditions, A 2B and A 3 receptors may be of relevance under pathological conditions. As an example, traumatic brain injury or cerebral ischemia lead to a massive surge in adenosine, elevating ambient levels of adenosine more than tenfold to levels sufficient to activate A 2B and A 3 receptors (Clark et al, 1997;Pearson et al, 2006). Recent findings suggest that the physiological consequences of A 3 receptor activation show dose-dependent effects: While CA1 hippocampal A 3 receptors stimulated by adenosine released during brief ischemia might exert A 1 receptor-like protective effects on neurotransmission, severe ischemia (i.e.…”