Evidence that the branched-chain alpha-keto acids accumulating in maple syrup urine disease induce morphological alterations and death in cultured astrocytes from rat cerebral cortex.

Severe neurological symptoms, cerebral edema, and atrophy are common features of the inherited metabolic disorder maple syrup urine disease (MSUD). However, the pathomechanisms involved in the neuropathology of this disease are not well established. In this study, we investigated the effects of the branched-chain keto acids (BCKA) alpha-ketoisocaproic (KIC), alpha-ketoisovaleric (KIV), and alpha-keto-beta-methylvaleric (KMV), which accumulate in MSUD, on astrocyte morphology and cytoskeleton reorganization. Cultured astrocytes from cerebral cortex of neonatal rats were exposed to various concentrations of the BCKA and cell morphology was studied. We observed that these cells changed their usual polygonal morphology when exposed to BCKA, leading to the appearance of fusiform or process-bearing cells. Furthermore, longer exposures to the BCKA elicited cell death at all concentrations studied, attaining massive death at the highest concentrations. Immunocytochemistry with anti-actin or anti-GFAP antibodies revealed that the BCKA induced reorganization of actin and GFAP cytoskeleton. In addition, astrocytes treated with lysophosphatidic acid, an upstream activator of the RhoA GTPase pathway, totally prevented the morphological alterations and cytoskeletal reorganization induced by KIV, indicating that this effect could be mediated by the RhoA signaling pathway. Furthermore, the effects of BCKA on astrocyte morphology were prevented by creatine. In addition, creatine kinase activity was inhibited by KIC and KIV; this inhibition was prevented by creatine, indicating that these keto acids compromise brain energy metabolism. Considering that astroglial cells are critical to brain development and functioning, it is conceivable that alterations of the actin network by BCKA may have important implications in astrocytic function and possibly in the pathogenesis of the neurological dysfunction and brain damage of MSUD patients. (c) 2004 Wiley-Liss, Inc.