[Histonet] RE: Perfusion technics for rodents

[gayle callis]

Several of our labs perfuse mice via heart (using a smaller gauge needle and
syringe filled with fixative, or with larger rodents, via a peristolic(sp?)
pump with PBS perfused first followed by a fixative. The animals are
anesthetized via an injection first, then perfused.  The perfusion itself is
euthanasia and with more adult animals, it is apparent when they are
euthanized/perfused when upper and lower legs extend and are stiff. With
really young animals, perfusion may be more difficult.   The perfusion
starts through the heart since it pumps at allow the perfusion to take
place.  If it is difficult to see a tiny heart and exact location to insert
a needle correctly, I suggest using safety glasses that are magnifiers.
These are available from various vendors.  With anesthetized hamsters or
mouse, the abdominal area is opened to expose the ascending/descending
aortas behind intestines. These blood vessels are severed to allow perfusion
fluids to flow away from the animal.     
 
As far as approval to anesthetize via injection (using an approved
anesthetic) and perfuse any animal, the university research laboratories
have strict rules to follow. An Animal Care Committee with sets up and
requires methods used that are guidelines from granting agencies (NIH, etc)
These rules are adhered to faithfully or a researcher can be severely
criticized, and probably with penalties in place.  They have regular
inspections to make sure they are doing these protocols correctly.   
 
I have watched both type of perfusions and the animal simply sleeps through
the procedure. It is a carefully orchestrated, painless and humane
protocol.There are some who may find this an unsavory topic, but I was
impressed at how humane it was compared to the old cervical dislocation
method to euthanize rodents.  That is no longer allowed.  
 
Gayle M. Callis
HTL(ASCP)HT,MT
Bozeman MT 
 
 
 
 
You Wrote:  
 
it probably would be better; but good luck getting approval to do 
that.
 
 
--On Tuesday, June 30, 2009 9:32 AM +0530 raghul 
<jayaprakash.raghul
<http://lists.utsouthwestern.edu/mailman/listinfo/histonet> <@t> rccltd.in>
wrote:
 
> Dear Dzung,
> 
> I wonder whether the perfusion would be good in an euthanized animal with
> the circulation arrested. It would be better to try it under deep
> anesthesia. Certainly perfusion would be better
> 
> Regards
> Raghul
> RCC laboratories India Pvt. Ltd
> 
> 
> Hi Everyone!!
> 
>    I am perfusing CO2 euthanized 3 weeks old mice with PBS only.  I am
> nicking the upper right atrium of heart to collect the gushed out blood
> and then perfusing through ventricle using a 21G butterfly needle and
> peristaltic pump.  Sometimes I see the lungs swelling up and fluid comes
> out of mouth.  Occasionally, I see the liver fade to light pink.  Most of
> the time the paws become white.  But the brain and spinal cord (my
> tissues of interest) are always white, and seem to have been perfused.  I
> was wondering how to improve this to get more consistent good perfusions,
> and what signs should I look for to indicate good perfusion?
> 
> Thanks much,
> 
> Dzung
> NIAID
>