Submitted on September 26, 2002
Accepted on February 27, 2003
24-h Langendorff-Perfused Neonatal Rat Heart to Study the Impact of Adenoviral Gene Transfer
Susanne Wiechert1, Ali El-Armouche2, Thomas Rau2, Wolfram-Hubertus Zimmermann2, and Thomas Eschenhagen3*
1 Institute of Experimental and Clinical Pharmacology, University of Erlangen-Nuremberg, Erlangen, Germany; Institute of Experimental and Clinical Pharmacology, University Hospital Hamburg-Eppendorf, Hamburg, Germany
2 Institute of Experimental and Clinical Pharmacology, University Hospital Hamburg-Eppendorf, Hamburg, Germany; Institute of Experimental and Clinical Pharmacology, University of Erlangen-Nuremberg, Erlangen, Germany
3 Institute of Experimental and Clinical Pharmacology, University Hospital Hamburg-Eppendorf, Hamburg, Germany
* To whom correspondence should be addressed. E-mail: t.eschenhagen{at}uke.uni-hamburg.de.
The human genome project has increased the demand for simple experimental systems that allow the impact of gene manipulations to be studied under controlled ex vivo conditions. We hypothesized that, in contrast to adult hearts, neonatal hearts allow longterm perfusion and efficient gene transfer ex vivo. A Langendorff perfusion system was modified to allow perfusion for >24 h with particular emphasis on uncompromised contractile activity, sterility, online measurement of force of contraction, inotropic response to 
-adrenergic stimulation, and efficient gene transfer. The hearts were perfused with serum-free medium (DMEM+M199, 4+1) supplemented with hydrocortison, triiodothyronine, ascorbic acid, insulin, pyruvate, L-carnitine, creatine, taurine, L-glutamine, mannitol, and antibiotics recirculating (500 ml/2 hearts) at 1 ml/min. Hearts from 2 day-old rats beat constantly at 135-155/min and developed active force of 1-2 mN. During 24 h of perfusion twitch tension increased to approximately 165% of initial values (p<0.05), whereas the inotropic response to isoprenaline remained constant. A decrease in total protein content of 10% and histological examination indicated moderate edema, but actin and calsequestrin concentration remained unchanged and perfusion pressure remained constant at 7-11 mmHg. Perfusion with a LacZ encoding adenovirus at 3x108 active virus particles yielded homogeneous transfection of about 80% throughout the heart and did not affect heart rate, force of contraction or response to isoprenaline when compared to uninfected controls (n=7 each). Taken together, the 24-h Langendorff-perfused neonatal rat heart is a relatively simple, inexpensive and robust new heart model that appears feasible as a test bed for functional genomics.
