A novel glutamate transporter blocker, LL-TBOA, attenuates ischaemic injury in the isolated, perfused rat heart despite low transporter levels.

Abstract

OBJECTIVES:

Loss of glutamate from cardiomyocytes during ischaemia may aggravate ischaemia-reperfusion injury in open heart surgery. This may be due to reversal of excitatory amino acid transporters (EAATs). However, the expression of such transporters in cardiomyocytes is ambiguous and quantitative data are lacking. Our objective was to study whether EAATs were expressed in the rat heart and to study whether blocking of transporter operation during cardiac ischaemia could be beneficial.

METHODS:

We used TaqMan real-time PCR and immunoisolation followed by western blotting to unequivocally identify EAAT subtypes in rat hearts. We used a novel high-affinity non-transportable competitive inhibitor, named LL-TBOA [(2S,3S)-3-(3-(6-(6-(2-(2-(2-(2-(2-aminoethoxy)ethoxy)-ethoxy)ethoxy) acetamido)hexanamido)- hexanamido)-5-(4-(trifluoromethyl)benzamido)benzyloxy) aspartic acid], to block EAAT-mediated transport during global ischaemia and reperfusion of isolated rat hearts.

RESULTS:

Rat hearts expressed EAAT subtypes 1 and 3, while subtypes 2 and 4 were not detected. Hearts were isolated and perfused with 1.6 µM LL-TBOA for 5 min before 30 min of induced global ischaemia and 60 min of reperfusion (n = 8). Control hearts were perfused either with the solvent dimethylsulfoxide 3.5 mM (n = 7) or with no pretreatment (n = 8). Infarct size was evaluated by triphenyl tetrazolium chloride (TTC) staining. LL-TBOA reduced infarct size from 33 ± 14 to 20 ± 5% (mean ± SD) (P = 0.015). Dimethylsulfoxide alone had no effect (35 ± 2%). Reperfusion arrhythmias were reduced by LL-TBOA (P = 0.009), but not by dimethylsulfoxide alone.

CONCLUSION:

Rat hearts express EAAT1 and EAAT3, but the mRNA levels are, respectively, ∼ 25 and 200 times lower than in the brain. Addition of LL-TBOA has a beneficial effect against ischaemia-reperfusion injury.

• Authors: Vladimir Martinov, Yvette Dehnes, Silvia Holmseth, Keiko Shimamoto, Niels Christian Danbolt, Guro Valen
• Journal: European journal of cardio-thoracic surgery : official journal of the European Association for Cardio-thoracic Surgery (Eur J Cardiothorac Surg) Vol. 45 Issue 4 Pg. 710-6 (Apr 2014) ISSN: 1873-734X [Electronic] Germany
• PMID: 24099732 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Amino Acid Transport System X-AG
• Excitatory Amino Acid Antagonists
• Aspartic Acid

Topics

• Amino Acid Transport System X-AG (antagonists & inhibitors)
• Animals
• Arrhythmias, Cardiac (drug therapy, prevention & control)
• Aspartic Acid (analogs & derivatives, chemistry, pharmacology, therapeutic use)
• Excitatory Amino Acid Antagonists (chemistry, pharmacology, therapeutic use)
• Heart Ventricles (drug effects)
• Myocardial Infarction (drug therapy)
• Myocardial Reperfusion Injury (drug therapy, pathology)
• Rats