Spreading Depolarization-Induced Adenosine Accumulation Reflects Metabolic Status In Vitro and In Vivo

Abstract

Spreading depolarization (SD), a pathologic feature of migraine, stroke and traumatic brain injury, is a propagating depolarization of neurons and glia causing profound metabolic demand. Adenosine, the low-energy metabolite of ATP, has been shown to be elevated after SD in brain slices and under conditions likely to trigger SD in vivo. The relationship between metabolic status and adenosine accumulation after SD was tested here, in brain slices and in vivo. In brain slices, metabolic impairment (assessed by nicotinamide adenine dinucleotide (phosphate) autofluorescence and O 2 availability) was associated with prolonged extracellular direct current (DC) shifts indicating delayed repolarization, and increased adenosine accumulation. In vivo, adenosine accumulation was observed after SD even in otherwise healthy mice. As in brain slices, in vivo adenosine accumulation correlated with DC shift duration and increased when DC shifts were prolonged by metabolic impairment (i.e., hypoglycemia or middle cerebral artery occlusion). A striking pattern of adenosine dynamics was observed during focal ischemic stroke, with nearly all the observed adenosine signals in the periinfarct region occurring in association with SDs. These findings suggest that adenosine accumulation could serve as a biomarker of SD incidence and severity, in a range of clinical conditions. Keywords: acute stroke; adenosine; brain slice; electrophysiology; energy metabolism; spreading depression INTRODUCTION Spreading depolarization (SD) of brain tissue is a self-propagating wave of neuronal and glial activation, carried by large transmembrane currents that depolarize cells and disrupt ionic gradients. 1,2 Spreading depolarization imposes a large metabolic burden on brain tissue, evidenced by depletion of energy substrates and O 2 3,4 and by accumulation of metabolic byproducts such as lactate and H + . 5,6 ATP concentration drops by nearly 50% during SD in normoxic conditions, 7 and metabolic depletion is expected to be more severe when occurring in tissue with limited substrate availability.Recent clinical studies strongly suggest that SD is involved in the pathophysiology of migraine with aura, thromboembolic stroke, traumatic brain injury, and subarachnoid hemorrhage. 8 While normoxic, normoglycemic SD can be noninjurious, 9 SD in the setting of metabolic compromise exacerbates neuronal injury. 10,11 Work in animal models has identified delayed repolarization, marked by prolonged extracellular direct current (DC) shifts, as a hallmark of injurious SD in metabolically compromised tissue. 10 Observational studies in human subjects have confirmed the association between prolonged DC shifts and poor outcome after SD, 12 but human studies have so far been limited by the requirement for invasive electrical recordings in patients with craniotomies. It would be helpful to establish additional biomarkers of metabolic status, to assess vulnerability to SD in injured brain.Extracellular adenosine accumulation can serve as a global indicator of energy charge....

Blockade of P2X7 Receptors or Pannexin-1 Channels Similarly Attenuates Postischemic Damage

“…32 Moreover, the effect that these antagonists have on MCAO, where their administration is 30 minutes after the occlusion, might indicate that P2X7 receptor or Panx1-channel inhibitors may reduce the propagation of spreading depression in the periinfarct area. 36 This finding together with our observations supports the idea that both P2X7r and Panx1 are recruited during brain ischemia as well, and that both channels constitute an amenable target for the prevention of neuronal death.…”

Spreading Depression, Spreading Depolarizations, and the Cerebral Vasculature

“…Depolarization also triggers Ca 2ϩ influx and a more than 10-fold drop in [Ca 2ϩ ] o (146,180,340), which, along with Na ϩ and water influx, leads to release of many if not all neurotransmitters and neuromodulators within the depolarized tissue. Extracellular glutamate, aspartate, glycine, GABA, and taurine concentrations increase during SD (117,232,326), and similar increases have been shown for adenosine, catecholamine, and ascorbate levels (277,278,325,391,465). The massive rise in [K ϩ ] o to levels sufficient to depolarize neighboring cells is the critical factor mediating the contiguous spread of the wave (i.e., reaction-diffusion model of SD propagation; FIGURE 2) (161,162,180).…”

“…Although some studies have failed to detect a significant change in ATP levels during SD (42,43,226,237,238,262), others have convincingly shown a decrease by as much as 50% (143,321,416), along with increased ADP and AMP (262). Adenosine levels are also elevated and correlate with the metabolic burden and mismatch imposed on the tissue by SD (278). The onset of these changes approximately coincides with the DC shift onset (143,321).…”

Evidence that adenosine contributes to Leao’s spreading depression in vivo

“…Initial observations were made in isoflurane-anesthetized mice (0.9-1.2% delivered by nose cone at 2 L/min, n ¼ 13 males, n ¼ 7 females) as previously described. 21,30 As isoflurane itself has CNS-depressant effects, 31 these initial findings were replicated and hypotheses further tested under urethane anesthesia (1.2-1.4 mg/g, i.p., n ¼ 25 males, n ¼ 13 females). Continuous systemic physiology was monitored non-invasively (Physiosuite, Kent Scientific Corporation, Torrington, CT) in urethane-anesthetized mice breathing room air, and values were stable over the course of experiments (pre-experiment heart rate 567.1 AE 2.1 bpm, spO 2 86.3 AE 0.2%, post-experiment heart rate 571.1 AE 2.4bpm, spO 2 87.0 AE 0.3%, n ¼ 36).…”