Biochimica et Biophysica Acta, 1097 (1991) 263-269 263 © 1991 Elsevier Science Publishers B.V. All rights reserved 0925-4439/91/$03.50 BBADIS 61088 Accumulation and excretion of long-chain acylcarnitine by rat hearts; studies with aminocarnitine Willem C. Hfilsmann l, Chris T.W.M. Schneijdenberg 2 and Arie J. Verkleij 2 l Thorax Centre, Erasmus University Rotterdam, Rotterdam (The Netherlands) and 2 Department of Molecular Cell Biology, State Unit,ersity Utrecht, Utrecht (The Netherlands) (Received 8 April 1991) Key words: Cardiac structure; Function; Acidosis; Ischemia; Long-chain acylcarnitine; Carnitine palmitoylcoenzyme A transferase During Langendorff perfusion of rat heart with aminocarnitine, long-chain acyicarnitine (LCAC) accumulates in heart cells, from which it is excreted by the heart. The heart function remains intact during this process. The accumulation of LCAC can be inhibited by the simultaneous addition of an inhibitor of the outer membrane carnitine paimitoylcoenzyme A transferase (CPT-1), indicating that aminocarnitine is a specific inhibitor of the inner membrane isoenzyme (CPT-2). LCAC accumulation is associated with glycogen depletion. After 60 min perfusion with aminocarnitine, electron microscopy shows large muitilameilar lipid vesicles, especially in cardiomy- ocytes, which are depleted in glycogen granula. Multilamellar lipid vesicles are also found in the bloodvessels. Extraction of the perfusate shows the presence of LCAC, fatty acid and phosphatidylethanolamine. Morphological analysis with freeze fracturing and thin sectioning furthermore reveals that the sarcolemma is not deteriorated during the export of LCAC to the coronary vessels. Since cardiac structures and functions are intact, LCAC alone is not the clue for ischemic damage. Therefore the present work supports the hypothesis that acidosis rather than LCAC is of primary importance to ischemic damage. Introduction Acidosis is probably the most important initiating factor in the loss of cardiac structure and function in ischemia [1-4]. Clustering of intramembranous parti- cles has been observed by freeze fracture studies [4,5]. Also, changes in the cytoskeleton of myocardial cells have been reported [6], as well as detachment of the cytoskeleton and loss of Ca 2+ from the inner leaflet of the sarcolemma [7]. These morphological changes may be based upon lateral phase separation and subsequent flip-flop of negatively charged phospholipid molecules from the inner to the outer leaflet. The destabilizing effect due to loss of bound calcium from negatively charged phospholipids by H + has been our preferred explanation for these phenomena. However, the local infiltration with endogenous detergent molecules such as long-chain acylcarnitine (LCAC), known to accumu- late in ischemia [8], may be an alternative or additional Correspondence: W.C. Hiilsmann, Thorax centre, Faculty of Medicine and Health Sciences, Erasmus University Rotterdam, P.O. Box 1738, 3000 DR Rotterdam, The Netherlands. destabilizing factor. In that light it may be noted that liposomal structures are formed within the mito- chondria after ischemia [5,9]. To investigate whether LCAC instead of acidosis could be the reason for sarcolemmal destabilization after ischemia, an experimental model was designed allowing the accumulation of LCAC in the absence of acidosis. In the present paper, we demonstrate that perfusion with L-aminocarnitine, a carnitine palmitoyl- coenzyme A transferase (CPT) inhibitor [10-12], is such a model. Injection of c-aminocarnitine into rats has been shown to cause accumulation of LCAC in a number of organs (Chiodi, P. and Maccari, F., personal communication), and in the liver this results in exces- sive acylation of cholesterol [13]. This suggests a pre- dominant inhibition of CPT-2, the mitochondrial inner membrane enzyme, while the formation of LCAC by CPT-1, the mitochondrial outermembrane enzyme, re- mains relatively intact. A higher efficiency of amino- carnitine as inhibitor of CPT-2 compared to CPT-1 has recently been reported [12]. Under our experimental setup of in vitro perfusion with aminocarnitine, CPT-1 activity was comparatively unaffected, as judged by the large accumulation of LCAC, as shown by biochemical