A new method for the investigation of capillary structure

Abstract

Numerous physiological conditions as well as behavioral conditions have been shown to influence central nervous system vascular structure. Many of the methods used to investigate these structural alterations take advantage of the visibility of viscous substances (e.g. India ink in gelatin) perfused into the vasculature. The high viscosity of the solution, however, can cause incomplete vessel perfusion. The aim of the present study was to test whether or not capillaries seen in tissue perfused with fixative, embedded in celloidin and stained with Methylene Blue–Azure II ( n=6) could be a useful alternative for the investigation of brain vascular structure. The method was compared to tissue from six rats perfused with India ink in gelatin and stained with cresyl violet. Qualitatively, vessels in the standard perfused tissue embedded in celloidin yielded clear vessels with stained pericytes. The two methods did not differ in branch point to cell ratio, length of individual capillaries, vessel length per mm 3, and capillary tortuosity. The capillary diameter was greater in the celloidin embedded tissue than in the India ink perfused tissue. Measuring the diameter between vessel walls appears to provide a more accurate measure than the widest distance between India ink pigments. Quantitative comparisons suggest that perfusion with standard fixative followed by embedding in celloidin provides vascular quantification comparable to that from India ink perfused tissue. The present method has several advantages, which include visualization of pericytes, increased probability of complete perfusion, clear view of cells that might otherwise be obscured by opaque vessels, and the possibility of using the alternate cerebral hemisphere for investigation of vascular ultrastructure.