Hepatic encephalopathy (HE) is an important cause of morbidity and mortality in patients with severe liver failure. Acute HE associated with fulminant hepatic failure has an extremely poor prognosis and specific therapy is not available, short of an emergency liver transplantation. Although its pathogenesis remains poorly understood, ammonia is strongly implicated as a neurotoxin, and astrocytes appear to be the primary target of ammonia neurotoxicity. Additionally, altered bioenergetics and oxidative stress are thought to play a major role in this disorder. These facts led to a consideration of the involvement of mitochondrial permeability transition (MPT) as a factor in the pathogenesis of HE and ammonia neurotoxicity. The MPT is a Ca2+-dependent, cyclosporin A (CsA)-sensitive process due to the opening of a pore in the inner mitochondrial membrane leading to a collapse of ionic gradients and ultimately to mitochondrial dysfunction. We have recently shown that ammonia induced the MPT in cultured astrocytes. We intend to examine the role of the MPT in HE and hyperammonemia using ammonia-treated neural cell cultures and in vivo models of HE/hyperammonemia (HA). Our working hypothesis is that ammonia induces the MPT in astrocytes, culminating in mitochondrial failure and astroglial dysfunction. A corollary of this concept is that inhibition or interference in the development of the MPT in astrocytes may ameliorate CNS dysfunction in HE.
The Specific Aims of this proposal are: 1) To identify the factors responsible for the ammonia-induced MPT in cultured neural cells. Our focus will be on agents implicated in the pathogenesis of HE/H that have also been shown to induce the MPT in other cells. Specifically, we will examine the role of Ca 2+, reactive oxygen species, nitric oxide, pH and glutamine. We will determine whether these factors are elevated in ammonia-treated cultures, and whether diminishing their production or blocking their actions reduces or abolishes the MPT. Additionally, we will examine possible sequential interrelationships among these factors. 2) To determine whether ammonia-induced abnormalities in astrocytes (morphological alterations, defects in neurotransmitter uptake, and cell swelling) are mediated by the MPT, we will investigate whether inhibitors of the MPT (CsA, bongkrekik acid) are capable of diminishing or blocking the deleterious effects of ammonia. 3) To investigate the involvement of mitochondrial dysfunction as a potential factor in MPT-mediated cell injury. We will determine the state of mitochondrial function after ammonia treatment, and then investigate whether improving energy metabolism will inhibit ammonia-induced cellular injury. 4) To clarify whether the MPT occurs in in vivo models of HE (thioacetarnide treatment) and hyperammonemia. We will also determine whether factors that inhibit the MPT in vitro (e.g., CsA, trifluoperazine) are also capable of doing so in vivo. Additionally, we will assess the ability of MPT blockers to improve the clinical, histopathologic, neurochemical abnormalities, and the extent of brain swelling observed in HE/HA. We believe that these studies will yield critical data bearing on the pathogenesis of HE, and may potentially aid in the development of novel therapeutic strategies for the treatment of this condition.

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National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
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Special Emphasis Panel (ZRG1-BDCN-3 (01))
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Doo, Edward
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University of Miami School of Medicine
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Jayakumar, A R; Bak, L K; Rama Rao, K V et al. (2016) Neuronal Cell Death Induced by Mechanical Percussion Trauma in Cultured Neurons is not Preceded by Alterations in Glucose, Lactate and Glutamine Metabolism. Neurochem Res 41:307-15
Rama Rao, Kakulavarapu V; Norenberg, Michael D (2014) Glutamine in the pathogenesis of hepatic encephalopathy: the trojan horse hypothesis revisited. Neurochem Res 39:593-8
Jayakumar, A R; Valdes, V; Tong, X Y et al. (2014) Sulfonylurea receptor 1 contributes to the astrocyte swelling and brain edema in acute liver failure. Transl Stroke Res 5:28-37
Jayakumar, Arumugam R; Tong, Xiao Y; Curtis, Kevin M et al. (2014) Decreased astrocytic thrombospondin-1 secretion after chronic ammonia treatment reduces the level of synaptic proteins: in vitro and in vivo studies. J Neurochem 131:333-47
Jayakumar, Arumugam R; Tong, Xiao Y; Curtis, Kevin M et al. (2014) Increased toll-like receptor 4 in cerebral endothelial cells contributes to the astrocyte swelling and brain edema in acute hepatic encephalopathy. J Neurochem 128:890-903
Rama Rao, Kakulavarapu V; Verkman, A S; Curtis, Kevin M et al. (2014) Aquaporin-4 deletion in mice reduces encephalopathy and brain edema in experimental acute liver failure. Neurobiol Dis 63:222-8
Jayakumar, A R; Norenberg, M D (2013) Endothelial-astrocytic interactions in acute liver failure. Metab Brain Dis 28:183-6
Jayakumar, A R; Ruiz-Cordero, R; Tong, X Y et al. (2013) Brain edema in acute liver failure: role of neurosteroids. Arch Biochem Biophys 536:171-5
Rao, Kakulavarapu V Rama; Brahmbhatt, Monica; Norenberg, Michael D (2013) Microglia contribute to ammonia-induced astrocyte swelling in culture. Metab Brain Dis 28:139-43
Jayakumar, A R; Tong, X Y; Ospel, J et al. (2012) Role of cerebral endothelial cells in the astrocyte swelling and brain edema associated with acute hepatic encephalopathy. Neuroscience 218:305-16

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