Measurement of capillary pulsations in the rat neocortex with two-photon laser scanning confocal microscopy
Authors: James P. McAllister, Mark E. Wagshul, Shams Rashid, Jie Li
Background
Hydrocephalus is associated with increased pulsations in the
cerebral aqueduct, as demonstrated by cine MRI, as well as
increased pulse pressure, as demonstrated by invasive intracranial
pressure monitoring. What has yet to be elucidated is the
relationship between increased pulsations and the pathophysiology
of hydrocephalus. Are increased pulsations an important
component of the pathophysiology, or simply an artefact of
decreased intracranial compliance? We have shown that under
normal circumstances, the transmission of arterial pulsations into
the cranium is minimized (the so-called Windkessel effect).
In this paper, we sought to demonstrate this effect directly by
measuring capillary pulsations with two-photon laser scanning
confocal microscopy.
Materials and Methods
Sprague-Dawley rats (4) were anaesthetized and a cranial window
was created. The dura was left intact and the craniotomy
sealed with a coverslip to maintain intracranial physiology.
Imaging was performed with a custom-built microscope (Olympus FV300
confocal microscope with a 1.5W Ti:Sapphire laser, externally
mounted Hamamatsu PMT's and an NA 0.9, 60x water immersion
objective). A fluorescent dye (70 KDa Dextran fluorescein)
was injected into the tail vein. Fluorescent vessels with
diameters from 5-15 m and depths of 50-300 m below the pial
surface were chosen. Flow was measured by repeatedly scanning
a vessel and observing the dark unlabeled red blood cells flowing
through the bright labelled plasma background. Flow
pulsatility was defined by the pulsatility index, i.e. peak to peak
flow (over each cardiac cycle) divided by mean velocity.
Results
Reliable flow waveforms were detected in approximately 100
vessels with a mean diameter of 10.96 ± 2.54 m, and at a mean
depth of 175.62 ± 56.58 m from the pial surface. Mean flow
velocity was 0.75 ± 0.55 mm/sec and mean pulsatility index was 21.2
± 13.2%. Data from ongoing experiments in hydrocephalic
animals will also be presented.
Conclusions
We have demonstrated the feasibility of measuring intracranial
capillary pulsatility within the neocortex of healthy rats.
These preliminary measurements show that pulsations are transmitted
from the macrovascular arterial flow into the
microvasculature. However, the amplitude of these flow
pulsations is small compared to the mean blood flow velocity.
As a comparison, pulsatility is commonly measured in humans within
the intracranial macrovasculature (e.g. MCA) using transcranial
ultrasound Doppler studies and is found to be 80-90% of the mean
flow. This technique will enable the study of changes in
capillary pulsatility in rat models of hydrocephalus and its
relationship to disease pathophysiology.
Department of Neurosurgery, Primary Children's Medical
Center, Salt Lake City, Utah
Email: pat.mcallister@hsc.utah.edu
