2.6.
VASCULAR PERFUSION
25
accomplish nothing worthwhile, and would run the risk of unnecessarily
damaging tissues.
There is doubt in my mind as to whether warm or cold fixing solution
is better. I am inclined to favor the latter, and we usually perfuse with
fixative recently removed from a refrigerator. It can be suggested that
cold fixative tends to keep vascular channels maximally open while at the
same time it slows down degenerative changes to allow more time for the
fixative to penetrate.
When the perfusion is regarded as complete and the emergent fluid is
only slightly pink, it seems a good practice to clamp off the exit port and
let a little more fixative flow into the vascular tree until some pressure
builds up on the venous side. Probably, also, it is desirable to hold this
pressure briefly, for perhaps 5 minutes. Thus it can be expected that the
capillary and venous bed will be fully dilated with a reserve quantity of
fixative. Then the cannula is removed and careful dissection of the tissue
of interest begins.
We have removed tissue from deep parts of the brain as well as from
the spinal cord 30 minutes or more after starting perfusion, and these
have proved to be well preserved. The retinal tissue illustrated in Fig. 3
was obtained in this way, and it may be noted that there was no separation of the sensory layers from the pigment layer as would be expected
ordinarily, and that the full thickness of the retina and choroid was well
preserved. In this way we have obtained good preservation of the interior
parts of the spinal cord including its gray matter and the surrounding
white matter. However, the peripheral white matter has not been as well
preserved. We have obtained well-preserved kidney cortex (Fig. 10),
although we have not been able to reach the medulla adequately by this
sort of vascular perfusion.
Unfortunately, it is quite impossible to perfuse as simply as this with
osmium tetroxide fixatives. There are at least two reasons for failure.
Decisive under most circumstances is the fact that osmium tetroxide acts
as an extreme vasoconstrictor so that the arteriolar bed ordinarily is
clamped shut as the first fixative reaches it. Little or no fixative usually
gets beyond this point in an arterial injection. This generally is true
even though efforts are made to counteract the vasoconstriction by procedures that can be expected to dilate. Thus, in our own laboratory we
have perfused with dissolved vasodilators before running in the fixative.
We have used gaseous vasodilators such as amyl nitrite. We have precooled animals so that their blood temperature was close to zero before
VASCULAR PERFUSION
25
accomplish nothing worthwhile, and would run the risk of unnecessarily
damaging tissues.
There is doubt in my mind as to whether warm or cold fixing solution
is better. I am inclined to favor the latter, and we usually perfuse with
fixative recently removed from a refrigerator. It can be suggested that
cold fixative tends to keep vascular channels maximally open while at the
same time it slows down degenerative changes to allow more time for the
fixative to penetrate.
When the perfusion is regarded as complete and the emergent fluid is
only slightly pink, it seems a good practice to clamp off the exit port and
let a little more fixative flow into the vascular tree until some pressure
builds up on the venous side. Probably, also, it is desirable to hold this
pressure briefly, for perhaps 5 minutes. Thus it can be expected that the
capillary and venous bed will be fully dilated with a reserve quantity of
fixative. Then the cannula is removed and careful dissection of the tissue
of interest begins.
We have removed tissue from deep parts of the brain as well as from
the spinal cord 30 minutes or more after starting perfusion, and these
have proved to be well preserved. The retinal tissue illustrated in Fig. 3
was obtained in this way, and it may be noted that there was no separation of the sensory layers from the pigment layer as would be expected
ordinarily, and that the full thickness of the retina and choroid was well
preserved. In this way we have obtained good preservation of the interior
parts of the spinal cord including its gray matter and the surrounding
white matter. However, the peripheral white matter has not been as well
preserved. We have obtained well-preserved kidney cortex (Fig. 10),
although we have not been able to reach the medulla adequately by this
sort of vascular perfusion.
Unfortunately, it is quite impossible to perfuse as simply as this with
osmium tetroxide fixatives. There are at least two reasons for failure.
Decisive under most circumstances is the fact that osmium tetroxide acts
as an extreme vasoconstrictor so that the arteriolar bed ordinarily is
clamped shut as the first fixative reaches it. Little or no fixative usually
gets beyond this point in an arterial injection. This generally is true
even though efforts are made to counteract the vasoconstriction by procedures that can be expected to dilate. Thus, in our own laboratory we
have perfused with dissolved vasodilators before running in the fixative.
We have used gaseous vasodilators such as amyl nitrite. We have precooled animals so that their blood temperature was close to zero before