Case example:

Center for Stroke Research Berlin, Charité University Medicine Berlin

Prof. Dr. med. Jens P. Dreier Charité University Medicine Berlin, Germany Center for Stroke Research Berlin

We discovered the phenomenon of inverse neurovascular coupling by coincidence in a rat model mimicking the conditions present at the boundary zone between subarachnoid blood and cortical surface after aneurysmal subarachnoid hemorrhage (aSAH).¹Dreier, J. P., K. Korner, N. Ebert, A. Gorner, I. Rubin, T. Back, U. Lindauer, T. Wolf, A. Villringer, K. M. Einhaupl, M. Lauritzen and U. Dirnagl (1998). “Nitric oxide scavenging by hemoglobin or nitric oxide synthase inhibition by N-nitro-L-arginine induces cortical spreading ischemia when K+ is increased in the subarachnoid space.” J Cereb Blood Flow Metab 18(9): 978-990. In this model, spreading ischemia resulted from topical application of artificial cerebrospinal fluid (ACSF) containing an elevated K⁺ concentration ([K⁺]_ACSF) in combination with either the nitric oxide synthase (NOS) inhibitor N^G-nitro-L-arginine (L-NNA) or the NO scavenger hemoglobin to the brain cortex (= NO↓/K⁺↑-model). Notably, the SD-induced spreading ischemia could be the sole cause of widespread cortical infarcts.²Dreier, J. P., N. Ebert, J. Priller, D. Megow, U. Lindauer, R. Klee, U. Reuter, Y. Imai, K. M. Einhaupl, I. Victorov and U. Dirnagl (2000). “Products of hemolysis in the subarachnoid space inducing spreading ischemia in the cortex and focal necrosis in rats: a model for delayed ischemic neurological deficits after subarachnoid hemorrhage?” J Neurosurg 93(4): 658-666. An NO donor caused spreading ischemia to revert to a normal neurovascular response to SD³Dreier, J. P., G. Petzold, K. Tille, U. Lindauer, G. Arnold, U. Heinemann, K. M. Einhaupl and U. Dirnagl (2001). “Ischaemia triggered by spreading neuronal activation is inhibited by vasodilators in rats.” J Physiol 531(Pt 2): 515-526. . A decade later, we observed spreading ischemia in aSAH patients for the first time.⁴Dreier, J. P., S. Major, A. Manning, J. Woitzik, C. Drenckhahn, J. Steinbrink, C. Tolias, A. I. Oliveira-Ferreira, M. Fabricius, J. A. Hartings, P. Vajkoczy, M. Lauritzen, U. Dirnagl, G. Bohner and A. J. Strong (2009). “Cortical spreading ischaemia is a novel process involved in ischaemic damage in patients with aneurysmal subarachnoid haemorrhage.” Brain 132(Pt 7): 1866-1881. In our recent clinical follow-up study, spreading ischemia occurred in the same time window in which serial neuroimaging-proven brain infarcts developed at the recording site.⁵Luckl, J., C. L. Lemale, V. Kola, V. Horst, U. Khojasteh, A. I. Oliveira-Ferreira, S. Major, M. K. L. Winkler, E. J. Kang, K. Schoknecht, P. Martus, J. A. Hartings, J. Woitzik and J. P. Dreier (2018). “The negative ultraslow potential, electrophysiological correlate of infarction in the human cortex.” Brain 141(6): 1734-1752. This confirmed our original translational hypothesis that spreading ischemia is involved in the pathogenesis of delayed ischemic stroke after aSAH.⁶Dreier, J. P., K. Korner, N. Ebert, A. Gorner, I. Rubin, T. Back, U. Lindauer, T. Wolf, A. Villringer, K. M. Einhaupl, M. Lauritzen and U. Dirnagl (1998). “Nitric oxide scavenging by hemoglobin or nitric oxide synthase inhibition by N-nitro-L-arginine induces cortical spreading ischemia when K+ is increased in the subarachnoid space.” J Cereb Blood Flow Metab 18(9): 978-990.

Following the recent concept of translational medicine by Marc Fisher⁷Fisher, M. (2013). “The spectrum of translational stroke research.” Neurol Res 35(5): 443-447., this development not only included classic translation from bench to bedside but also reverse and lateral translation. Lateral translation occurs when a proven clinical therapy or technology stimulates basic and/or clinical researchers to enhance these proven modalities. Reverse translation occurs when clinical advances encourage basic science researchers to understand more about the mechanisms underlying clinical observations, technologies or therapies. For example, driven by our clinical findings, we have continued to elucidate the mechanism underlying spreading ischemia in rodent experiments.

One of our recent experimental studies has recently suggested that prolonged elevation of the baseline extracellular K⁺ concentration ([K⁺]_o) causes rundown of the α₂ Na⁺/K⁺-ATPase, located in a microdomain above the endoplasmic reticulum.⁸Major, S., G. C. Petzold, C. Reiffurth, O. Windmuller, M. Foddis, U. Lindauer, E. J. Kang and J. P. Dreier (2017). “A role of the sodium pump in spreading ischemia in rats.” J Cereb Blood Flow Metab 37(5): 1687-1705. This may result in increased Ca²⁺ uptake by internal stores of astrocytes, vascular myocytes and pericytes since Ca²⁺ outflux via plasmalemmal Na⁺/Ca²⁺-exchanger declines. Augmented Ca²⁺ mobilization from internal stores during SD could then enhance vasoconstriction, thereby contributing to spreading ischemia. However, augmented Ca²⁺ mobilization activates constitutive NOS, thereby antagonizing vasoconstriction. This could explain why spreading ischemia in the NO↓/K⁺↑-model requires reduced NO availability in addition to elevated baseline [K⁺]_o. Further insight into the mechanisms underlying inverse neurovascular coupling could help us to find novel treatment strategies in the clinic following the tenet espoused by Max Planck that ‘insight must precede application’.

The inverse neurovascular response to SD is observed as a spreading perfusion deficit causing prolongation of SD (Dreier 2011). Laser Speckle Contrast Analysis (LASCA or LSCI) is an ideal tool to differentiate the normal (fig. 1) from the inverse neurovascular response to SD (fig. 2) because it visualizes the perfusion changes in space and time, and is easily combined with various electrophysiological methods to measure, for example, SD or additional variables such as tissue partial pressure of oxygen.

Figure 1

Figure 2

Open cranial window experiments in rats. Using Perimed’s PeriCam PSI laser speckle contrast analysis imager, two-dimensional maps of cortical cerebral blood flow are obtained with very high spatial and temporal resolution. Regions of interest can be defined (circles) in which cerebral blood flow is quantified over time. Figure 1 shows typical perfusion changes of an SD-induced spreading hyperemia and figure 2 of an SD-induced spreading ischemia. The left panels give consecutive images and the right panels the time courses of cerebral blood flow in the regions of interest. Click on the images for a full video.