2.6.
VASCULAR PERFUSION
27
heart is amputated and artificial respiration discontinued. A glass cannula, fashioned with a bulbous tip, is inserted quickly into the left
ventricle, through the aortic valve, and tied into place at the root of the
aorta with the previously placed ligature.
The perfusion is begun with 6-7 ml of balanced salt solution, and
immediately followed with 100-150 ml of Palade's buffered osmium
tetroxide (Chapter 3.2) to which 5.4 mg of anhydrous CaCl 2 /ml of
final solution is added. Both the saline solution and the initial portion of
the fixative solution are warmed by means of heating tapes wound
around the rubber tubing of the perfusion apparatus. The temperature of
the heating tapes is adjusted so the rubber tubing becomes warm but not
hot to the touch (approximately 30°C). The bulk of the fixing solution
in the reservoir is chilled to ice temperature. Perfusion is carried out
under a head of pressure equivalent to 5 ft of water. After the initial
flow washes the blood out of the vascular tree, and osmium tetroxide is
detectable in the return pouring out of the open right ventricle, the
heating tapes are disconnected, and the flow is reduced to a slow trickle
so that 20-40 minutes are required to pass the total of 150 ml of cold
fixative through the head end of the carcass. The entire operative procedure, from the opening of the chest to the beginning of perfusion,
need take only 2-3 minutes. Less than 1 minute need elapse from the
cessation of respiration until the first sign of fixation (blackening of nose
and ears).
When these authors are interested in preserving the lower spinal cord
by perfusion through the heart, they first clamp the iliac, mesenteric,
and carotid arteries in that sequence. Alternatively, they use a retrograde
aortic perfusion from the point of the aortic bifurcation after clamping
the aortic arch. In this instance they do not clamp the mesenteric arteries
and as a result achieve an incomplete perfusion of abdominal viscera.
There are also limited anatomical situations where retrograde injections through venous channels make perfusion with osmium tetroxide
fixative relatively easy and reliable. Liver and spleen are obvious possibilities. Excellent micrographs of liver cells preserved by vascular
perfusion have in fact been published. Of course other natural body
cavities, such as the cerebral spinal fluid passage-ways, can be used as
routes for the in situ preservation of neighboring tissues. The epithelial
linings of tubular organs can be similarly reached with fixative, perhaps
after vigorous flushing with a balanced salt solution (Chapter 2.4).
The cost of osmium tetroxide, as well as dangers inherent in handling
VASCULAR PERFUSION
27
heart is amputated and artificial respiration discontinued. A glass cannula, fashioned with a bulbous tip, is inserted quickly into the left
ventricle, through the aortic valve, and tied into place at the root of the
aorta with the previously placed ligature.
The perfusion is begun with 6-7 ml of balanced salt solution, and
immediately followed with 100-150 ml of Palade's buffered osmium
tetroxide (Chapter 3.2) to which 5.4 mg of anhydrous CaCl 2 /ml of
final solution is added. Both the saline solution and the initial portion of
the fixative solution are warmed by means of heating tapes wound
around the rubber tubing of the perfusion apparatus. The temperature of
the heating tapes is adjusted so the rubber tubing becomes warm but not
hot to the touch (approximately 30°C). The bulk of the fixing solution
in the reservoir is chilled to ice temperature. Perfusion is carried out
under a head of pressure equivalent to 5 ft of water. After the initial
flow washes the blood out of the vascular tree, and osmium tetroxide is
detectable in the return pouring out of the open right ventricle, the
heating tapes are disconnected, and the flow is reduced to a slow trickle
so that 20-40 minutes are required to pass the total of 150 ml of cold
fixative through the head end of the carcass. The entire operative procedure, from the opening of the chest to the beginning of perfusion,
need take only 2-3 minutes. Less than 1 minute need elapse from the
cessation of respiration until the first sign of fixation (blackening of nose
and ears).
When these authors are interested in preserving the lower spinal cord
by perfusion through the heart, they first clamp the iliac, mesenteric,
and carotid arteries in that sequence. Alternatively, they use a retrograde
aortic perfusion from the point of the aortic bifurcation after clamping
the aortic arch. In this instance they do not clamp the mesenteric arteries
and as a result achieve an incomplete perfusion of abdominal viscera.
There are also limited anatomical situations where retrograde injections through venous channels make perfusion with osmium tetroxide
fixative relatively easy and reliable. Liver and spleen are obvious possibilities. Excellent micrographs of liver cells preserved by vascular
perfusion have in fact been published. Of course other natural body
cavities, such as the cerebral spinal fluid passage-ways, can be used as
routes for the in situ preservation of neighboring tissues. The epithelial
linings of tubular organs can be similarly reached with fixative, perhaps
after vigorous flushing with a balanced salt solution (Chapter 2.4).
The cost of osmium tetroxide, as well as dangers inherent in handling