Elsevier

Brain Research

Volume 1111, Issue 1, 21 September 2006, Pages 111-116
Brain Research

Research Report
NADPH-oxidase activity is elevated in penumbral and non-ischemic cerebral arteries following stroke

https://doi.org/10.1016/j.brainres.2006.06.082Get rights and content

Abstract

Reactive oxygen species play a role in neuronal damage following cerebral ischemia–reperfusion. We tested whether activity of the superoxide-generating enzyme, NADPH-oxidase, is enhanced in cerebral arteries within, adjacent and distant from the ischemic core. The right middle cerebral artery (MCA) of conscious rats was temporarily occluded by perivascular injection of endothelin-1 to induce stroke (ET-1; n = 19). Control rats were injected with saline (n = 9). At 24 h or 72 h post-administration of ET-1, the MCA and its branches within the ipsilateral penumbra and infarcted core, corresponding arteries in the contralateral hemisphere, and basilar artery were excised. Anatomically similar arteries were excised from saline-injected rats. At 24 h after stroke, NADPH-stimulated superoxide production by arteries from the infarcted core did not differ from levels generated by arteries from control rats, whereas levels were significantly lower 72 h after stroke. However, at both time points after stroke, superoxide production by arteries from the ischemic penumbra was 8-fold greater than levels generated by arteries from control rats. Surprisingly, even in the non-ischemic arteries from the contralateral hemisphere and in the basilar artery, superoxide production was increased ∼ 4- to 6-fold at 24 h, but had returned to normal 72 h after stroke. The NADPH-oxidase inhibitor, diphenyleneiodonium, virtually abolished superoxide production by all arteries. Thus, the activity of NADPH-oxidase is enhanced in cerebral arteries from the ischemic penumbra at 24 h and 72 h following cerebral ischemia. Additionally, NADPH-oxidase activity is temporarily enhanced after cerebral ischemia within arteries from non-ischemic parts of the brain.

Introduction

A reduction in cerebral blood flow to less than 10% of normal results in irreversible death to affected neurons. Between this infarct core and unaffected regions of the brain lies an area of constrained blood flow, referred to as the penumbra. Neurons within the penumbra die over a more protracted period, which may extend from hours to days (Dirnagl et al., 1999, Fisher and Garcia, 1996).

During reperfusion, the function and integrity of cerebral arteries are critical to support cerebral blood flow and thus minimize further neuronal injury (Fagan et al., 2004). A growing body of evidence suggests that reactive oxygen species (ROS), such as superoxide (O2), contribute to neuronal damage during the early phases of reperfusion (Chan, 2001). Indeed, excessive ROS production in cerebral arteries during the first few hours of reperfusion may result in further vascular dysfunction and a decline in perfusion of the previously ischemic brain (Kontos et al., 1992, Nelson et al., 1992). It has been suggested that ROS may also participate in delayed neuronal injury in the penumbra (Dirnagl et al., 1999). However, it is unclear whether ROS production in cerebral arteries is elevated at such later time points. NADPH-oxidases are now thought to be the primary generators of O2 within the vasculature (Ellmark et al., 2005, Griendling et al., 1994, Miller et al., 2005). Thus, the aim of this study was to test whether NADPH-oxidase activity and ROS production within arteries of ischemic and adjacent brain regions is altered following stroke induced by temporary middle cerebral artery (MCA) occlusion.

Section snippets

Outcome of ET-1 or saline administration

All rats displayed no signs of neurological deficit pre-surgery, but had low deficit scores prior to ET-1/saline administration (0 h, Figs. 1A, B). This slight deficit was presumably as a result of unavoidable neuronal damage during cannula implantation. Control rats showed no signs of stroke. In contrast, rats displayed neurological deficits, indicative of stroke, within 10 min of injection of ET-1 (data not shown). 24 h and 72 h after administration of ET-1 rats exhibited a greater deficit

Discussion

The major new finding of this study is that NADPH-oxidase activity in cerebral arteries from the ischemic penumbra is elevated for up to 3 days after mild stroke induced by temporary MCA occlusion in rats. A second, unexpected finding is that for the first 24 h after stroke, the increase in vascular NADPH-oxidase activity was not confined to penumbral arteries but also occurred in arteries from the contralateral (non-ischemic) hemisphere and in the basilar artery.

In the present study, we

Surgical preparation

All procedures were approved by the institutional animal ethics committee. Male hooded Wistar rats (n = 31; weight, 280–320 g) were anesthetized with sodium pentobarbital (60 mg/kg, i.p.). A guide cannula was stereotaxically implanted into the piriform cortex 2 mm dorsal to the ipsilateral MCA as previously described (Sharkey et al., 1993). The stereotaxic coordinates were modified for this rat strain (0.2 mm anterior, − 5.2 mm lateral and − 6.1 ventral, according to a stereotaxic atlas) (Paxinos

Acknowledgments

These studies were supported by an Institute Block Grant (No. 983001) from the National Health and Medical Research Council of Australia. C.G.S. is a Senior Research Fellow of the National Health and Medical Research Council of Australia.

References (33)

  • P.H. Chan

    Reactive oxygen radicals in signaling and damage in the ischemic brain

    J. Cereb. Blood Flow Metab.

    (2001)
  • S.Y. Cheranov et al.

    TNF-α dilates cerebral arteries via NAD(P)H oxidase-dependent Ca2+ spark activation

    Am. J. Physiol.: Cell Physiol.

    (2006)
  • G.W. De Keulenaer et al.

    Tumour necrosis factor alpha activates a p22phox-based NADH oxidase in vascular smooth muscle

    Biochem. J.

    (1998)
  • M. De Ryck et al.

    Photochemical stroke model: flunarizine prevents sensorimotor deficits after neocortical infarcts in rats

    Stroke

    (1989)
  • S.P. Didion et al.

    Effects of NADH and NADPH on superoxide levels and cerebral vascular tone

    Am. J. Physiol.: Heart Circ. Physiol.

    (2002)
  • S.H. Ellmark et al.

    The contribution of Nox4 to NADPH oxidase activity in mouse vascular smooth muscle

    Cardiovasc. Res.

    (2005)
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