NO synthase inhibition and elevated extracellular K+ produce cortical spreading ischemia

Jens P. Dreier, Tobias Back, Katrin Körner, Astrid Görner, Ulrich Dirnagl

Introduction

Cortical spreading depression (CSD) is a depolarization wave moving over the brain cortex with a speed of 3 to 5 mm/min. CSD is accompanied by large changes of the ionic homeostasis. Restoration of the changes requires energy. The energy supply is increased via increased regional cerebral blood flow (rCBF). The coupling between the increased metabolic workload and increased rCBF during CSD is not understood. One candidate to mediate coupling during CSD is nitric oxide (NO). In 2 studies inhibition of the NO producing enzyme NO synthase (NOS) resulted in a CSD induced initial hypoperfusion (hyporCBF) (Duckrow 1993 Brain Res 618:190-195, Fabricius et al 1995 Am J Physiol 269 in press). We have studied whether hyporCBF depends on the increase of the extracellular K+ concentration ([K+]o) during CSD.

Methods

58 male Wistar rats were anesthetized with thiopental-sodium. A closed cranial window was implanted over the right parietal cortex. The dura was cut permitting continuous superfusion of artificial cerebrospinal fluid (ACSF). The NOS-inhibitor N(-nitro-L-arginine (L-NA, 1 mmol/L) and the NO-donor S-nitroso-N-acetylpenicillamine (SNAP, 500 µmol/L) were solved in ACSF and applied topically. Elevation of the cerebrospinal K+ concentration ([K+]ACSF) from 3 to 20 mmol/L was balanced by reduction of the cerebrospinal Na+ concentration. RCBF was measured with laser-Doppler flowmetry and calculated comparing relative changes.The DC-potential was measured under the cranial window with a cortical surface electrode. CSD was elicited from a second open cranial window 5 mm apart from the closed cranial window.

Results

Our key findings are shown in fig. 1. A, B and C are 3 different experiments

• Fig. 1Aa, Ba and Ca: Typical rCBF and DC-potential responses to CSD under control conditions. The most prominent feature of rCBFCSD is the large increase (rCBFspike). The most prominent feature of DCCSD is the negative shift (negDCCSD)
• Fig. 1Ab, Bb and Cb: CSD responses 2 hours after superfusion with L-NA. Reduction of rCBFspike but no other changes. HyporCBF was induced in only ¼ of the animals
• Fig. 1Ac, Bc and Cc: CSD responses 1 hour later after superfusion of [K+]ACSF = 20 mmol/L during continued NOS inhibition. In all animals hyporCBF was induced. Contrary to other studies hyporCBF was prolonged, more pronounced and associated with a large increase and elongation of negDCCSD. In fact, the minimum level of rCBF during hyporCBF in fig. 1Cc is 12% compared to baseline, an ischemic level. RCBF remains below 30% for more than 20 minutes. On average the minimum level of rCBF during hyporCBF was 33 ( 21%. The duration of hyporCBF was 1063 ( 1534 s
• Fig. 1Bd: The changes of the CSD responses are reversible with SNAP

We also found that no hyporCBF is induced by elevation of [K+]ACSF to 20 mmol/L without NOS inhibition but the amplitude of negDCCSD increased.

Discussion

Since hyporCBF can reach ischemic levels, the spreading is preserved and the DC-potential changes are reminiscent of periinfarct depolarizations indicating an energy deficit, the CSD derivative under NOS inhibition and elevated [K+]ACSF is refered to as cortical spreading ischemia (CSI).

Since NOS inhibition is a stable condition, the acutely occuring hyporCBF requires a second factor that constricts the vessels on top of NOS inhibition. This second factor may be [K+]o, since (1) the peak of [K+]o accompanying CSD occurs before and during the initial phase of rCBFCSD, (2) [K+]o increases during CSD to levels between 30 and 80 mmol/L where it acts as a vasoconstrictor, (3) a consistent interaction of [K+]o and NO is described, (4) elevation of [K+]ACSF that promotes the occurence of hyporCBF is likely to increase the peak of [K+]o during CSD as indicated by the increase of negDCCSD both with and without NOS inhibition (the DC-potential reflects [K+]o, see e.g. Richter et al. (1993) Neurosci Lett 152:65-68), (5) the elongation of rCBFCSD is accompanied by elongation of negDCCSD reflecting [K+]o (a possible explanation may be that hyporCBF reduces the energy supply during the increased metabolic workload of CSD; ATP should fall consequently; the Na+-K+-ATPase should fail to pump K+ ions back into the cells; [K+]o remains high; vasoconstriction persists and a vicious circle of prolonged ischemia is induced accompanied by prolonged increase of [K+]o as indicated by prolonged negDCCSD).

CSI is a new phenomenon of the cerebrovasculature, possibly a new mechanism related to ischemia. Contrary to other mechanisms of ischemia such as thrombosis, embolism and vasospasm the ischemia of CSI is not triggered by the vessels but by neuronal and glial cells that secondarily induce vasoconstriction on the basis of functionally changed vessels.

CSI may be a clinically relevant pathomechanism e.g. in migrainous stroke or in the delayed ischemic deficits after subarachnoid hemorrhage which is a condition with protracted increase of [K+]o and protracted critical reduction of the cGMP levels in the vascular walls possibly by the selective NO antagonist hemoglobin. Only recently a rCBF response resembling CSI combined with a DC-potential and [K+]o shift characteristic of CSD was measured in human cortex in a patient with severe head trauma accompanied by cerebral hemorrhage (Mayevsky et al. (1995) J Cereb Blood Flow Metab 15 (Suppl 1): S34).

Publications

1. Dreier JP, Körner K, Görner A, Lindauer U, Weih M, Villringer A, Dirnagl U (1995) Nitric oxide modulates the CBF response to increased extracellular potassium J Cereb Blood Flow Metab 15:914-191, 1995
2. Dreier JP, Görner A, Körner K, Lindauer U, Back T, Dirnagl U (1995) Nitric oxide modulates the CBF response to spreading depression J Cereb Blood Flow Metab 15 (Suppl 1):S155
3. Dreier JP, Görner A, Körner K, Back T, Lindauer U, Wolf T, Villringer A, Lauritzen M, Dirnagl U NOS inhibition and elevated potassium produce cortical spreading ischemia submitted
4. Dreier, J.P., Ebert, N., Reuter, U., Wolf, T., Leistner, S., Villringer, A. and Dirnagl, U. Pharmacological transformation of spreading depression (SD) related hyperperfusion to an acute and longlasting hypoperfusion by NOS-inhibiton and ouabain, Soc. Neurosci. Abstr. 23 (1997) (in press)(Abstract)