Anna Lena Keller

Anatomical investigations of the cerebral microvascular system and the relationship of vascular density and local oxidative metabolism

Anna Lena Keller and external collaborators Johannes Reichold* and Bruno Weber*

(*University and ETH Zürich)

Introduction

Many functional imaging techniques, including fMRI (functional magnetic resonance imaging) are based on the local cerebral blood flow (CBF) increase that occurs concurrently with increases in neuronal activity. The structural properties of the brain’s microvasculature at the basis of this neurovascular coupling are still not well understood. We have developed a range of methods to investigate the cerebrovascular network in all its details.

Goals

The network properties of the cerebral vasculature are not known in detail, albeit this information is of utmost importance to improve the comprehension of the regulatory mechanisms of CBF and the interpretation of hemodynamic signals. Our recent emphasis lies on the processing of 3-dimensional data to extract reliable parameters for the modeling of blood flow dynamics.

Methods

Fluorescent Immunohistochemistry: On brain sections from macaque and squirrel monkeys we fluorescently labeled the blood vessels with an antibody against Collagen type IV. COX Staining: On some of the immunohistochemically labeled sections we visualized the activity of cytochrome oxidase, a marker of the oxidative metabolism, and correlated it with the vascular density. Corrosion Cast Technique: We injected two Macaca nemestrina with a methyl methacrylate resin. After removal of all brain tissues the casts were imaged with a scanning electron microscope (SEM). Synchrotron-radiation Based X-ray Microscopy (srXTM): A dispersed suspension of radio opaque BaSO4 was injected in the brains of rats and macaques and absorption-based srXTM was performed with a spatial resolution of 700 nm in a volume of ~1.5 mm³.

Initial results

The immunohistochemical approach resulted in a detailed quantification of vascular densities in layers and areas of the visual cortex [1]. Included as well are first estimates of an arterio-venous ratio derived from corrosion cast preparations. A tight correlation of vascular density and oxidative metabolism in several regions of V1 was revealed by immunolabeling of COX stained sections [2]. The first results of the srXTM approach lead to preliminary 3-dimensional representations of the vasculature, enabling the simulation of changes in cortical blood flow (CBF) [3].

Initial conclusion

The methodological portfolio we developed allows for an exact qualitative and quantitative assessment of the cerebral vasculature and can be applied to any brain region of interest. With more recently gained srXTM data of higher quality and a newly established method to define arteries and veins within the datasets, the existing vascular network graph can be refined which is a requirement for a more reliable modeling of CBF.

References

1.      Weber B., Keller AL., Reichold J., Logothetis NK. (2008) The microvascular system of the striate and extrastriate visual cortex of the macaque, Cerebral Cortex 18 2318-2330.

2.      Keller AL., Schüz A., Logothetis NK., Weber B. (2011) Vascularization of cytochrome oxidase-rich blobs in the primary visual cortex of squirrel and macaque monkeys The Journal of Neuroscience 31 1246-1253.

3.      Reichold J., Stampanoni M., Keller AL., Buck A., Jenny P., Weber B (2009) Vascular graph model to simulate the cerebral blood flow in realistic vascular networks, Journal of Cerebral Blood Flow & Metabolism 29 1249-1443.

Figures

Figure 1:

Scanning electron micrograph of a vascular corrosion cast preparation from the macaque monkey striate cortex. It shows a view across all cortical layers, covering approximately 1.7 mm from surface to white matter, larger arteries and veins shaded red and blue. Automated mosaic-like scanning enables the investigation of large specimen, though with the superior spatial resolution of the scanning elect ron microscope.

Figure 2:

Cerebrovascular networks obtained from synchrotron-radiation based x-ray tomography. The left image depicts a vascular network graph from the rat somatosensory cortex. To the right, a separated arterial tree from the graph on the left is shown. By extracting the larger vessels from the network, differences in the morphology of arteries and veins can be analyzed. Such representations as above and the parameters one can derive from them are used for fluid dynamics modeling of CBF.