Cranial Window Assessments in Experimental aSAH in Mice - Animal Models of Acute Neurological Injuries: Injury and Mechanistic Assessments - page 486

6. Drilling of a circular bone fl ap (diameter approx. 6 mm),
overlapping the superior sagittal sinus under continuous
irrigation and cleaning to avoid damage to the brain by heat or
direct injury and to provide clear vision.
7. Special attention should be paid to the drilling over the sagittal
suture, where the sinus is adherent to the bone.
8. Removal of the bone fl ap should be performed under highest
magnifi cation to avoid damage to the sinus or bridging veins.
9. After removal of the bone fl ap cleaning is facilitated by irriga-
tion and careful use of gauze.
10. For dural opening and removal an intracutaneous needle can
be used. The opening should be achieved at the far lateral
region of the craniectomy.
11. Bleedings should be stopped by irrigation only.
12. Be aware that the brain surface is to be kept moist at all times.
13. The window glass coverslip is threaded into position and fi xed
with glue. No air should be trapped beneath the glass slide.
14. Skin closure is achieved by primary suture at the end of the
procedure.
15. Note that for assessment of infl ammatory changes the fi rst
measurement should be performed at least 2 days after implan-
tation of the window to eliminate the infl uence of the operative
trauma.

Fluorescence microscopy is a widely used standard observation and
evaluation technique for a variety of “windows” to the vasculature.
It uses light of a certain wavelength to illuminate specifi cally applied
fl uorophores (e.g. green fl uorescent protein, fl uorescein, etc.).
A fi lter fi lters the excitation light from the fl uorescence light, thus giv-
ing a 2D fl uorescent image with an improved signal-to-noise ratio.
Intravital fl uorescence microscopy allows for analysis of the
following parameters:

3.2. Fluorescence
Microscopy

1. Vascular length ( L ) and diameter (Dv).

2. Luminal surface area ( A = Dv × L × p ).

3. Mean blood fl ow velocity ( V blood = V max /(2-( L /Dv)²), where V max
represents the maximum leukocyte velocity.

4. Wall shear rate ( g = V blood × 8/Dv).

5. Wall shear stress ( t = g × 0.025) 0.025 is assumed to be the blood
viscosity under normal conditions.

6. Total cellular fl ux (TF = number of cells passing a distinct vessel
per observation period).

7. Rolling fl ux (RF = number of rolling cells per observation period).

8. Sticking fl ux (SF = number of stationary cells >20 s).