Gabapentin Attenuates Oxidative Stress and Apoptosis in the Diabetic Rat Retina.

Neurodegeneration in diabetic retina has been widely considered as initiating factor that may lead to vascular damage, the classical hallmark of diabetic retinopathy. Diabetes induced altered glutamate metabolism in the retina, especially through glutamate excitotoxicity might play a major role in the neurodegeneration. Increased level of branched chain amino acids (BCAAs) measured in diabetic retina might cause an increase in the neurotoxic level of glutamate by transamination of citric acid cycle intermediates.

Molecular basis for increased lactate formation in the Müller glial cells of retina.

Müller glial cells are highly metabolic active cells that compensate for the high energy demand of retinal neurons. It has been believed that glucose provides the energy needs by the complete oxidation within Müller cells. However, numerous studies indicated that glial cells convert the majority of glucose to lactate, which may serve as an energy source for neurons.

Regulation of glutamate metabolism by hydrocortisone and branched chain keto acids in cultured rat retinal Müller cells (TR-MUL).

Glutamate released from retinal neurons during neurotransmission is taken up by retinal Müller cells, where much of the amino acid is subsequently amidated to glutamine or transaminated to α-ketoglutarate for oxidation. Müller cell glutamate levels may have to be carefully maintained at fairly low concentrations to avoid excesses of glutamate in extracellular spaces of the retina that would otherwise cause excitotoxicity. We employed a cultured rat retinal Müller cell line in order to study the metabolism and the role of Müller cell specific enzymes on the glutamate disposal pathways.

Influence of insulin on glutamine synthetase in the Müller glial cells of retina.

Glutamine synthetase (GS), a Müller cell specific enzyme in the retina, is the key enzyme involve in glutamate metabolism. The goal of this study was to investigate the expression and regulation of GS by insulin in the cultured rat retinal Müller cells. Immunocytochemical and immunoblotting experiments showed that the cultured Müller cells express GS protein under normal cell culture conditions.

The influence of diabetes on glutamate metabolism in retinas.

Excised retinas from euglycemic and diabetic Sprague-Dawley rats were studied to evaluate differences in glutamate metabolism related to diabetes. Reports suggest, neuronal cell death possibly caused by glutamate excitotoxicity, is an early consequence of diabetes. To monitor the influence of diabetes on glutamate metabolism, we measured glutamatergic neurotransmission, anaplerotic glutamate synthesis from (14) CO(2) and pyruvate as well as rates of glutamate cataplerosis ([U-(14) C]glutamate to (14) CO(2) and (14) C-pyruvate).

Links between insulin resistance, adenosine A2B receptors, and inflammatory markers in mice and humans.

Objective: To determine the mechanisms by which blockade of adenosine A(2B) receptors (A(2B)Rs) reduces insulin resistance.

Research Design And Methods: We investigated the effects of deleting or blocking the A(2B)R on insulin sensitivity using glucose tolerance tests (GTTs) and hyperinsulinemic-euglycemic clamps in mouse models of type 2 diabetes. The effects of diabetes on A(2B)R transcription and signaling were measured in human and mouse macrophages and mouse endothelial cells.

Cardiolipin remodeling by ALCAT1 links oxidative stress and mitochondrial dysfunction to obesity.

Oxidative stress causes mitochondrial dysfunction and metabolic complications through unknown mechanisms. Cardiolipin (CL) is a key mitochondrial phospholipid required for oxidative phosphorylation. Oxidative damage to CL from pathological remodeling is implicated in the etiology of mitochondrial dysfunction commonly associated with diabetes, obesity, and other metabolic diseases.

A multistep validation process of biomarkers for preclinical drug development.

Biomarkers that can be measured in preclinical models in a high-throughput, reproducible manner offer the potential to increase the speed and efficacy of drug development. Development of therapeutic agents for many conditions is hampered by the limited number of validated preclinical biomarkers available to gauge pharmacoefficacy and disease progression, but the validation process for preclinical biomarkers has received limited attention. This report defines a five-step preclinical biomarker validation process and applies the process to a case study of diabetic retinopathy.

Dual mechanism of brain injury and novel treatment strategy in maple syrup urine disease.

Maple syrup urine disease (MSUD) is an inherited disorder of branched-chain amino acid metabolism presenting with life-threatening cerebral oedema and dysmyelination in affected individuals. Treatment requires life-long dietary restriction and monitoring of branched-chain amino acids to avoid brain injury. Despite careful management, children commonly suffer metabolic decompensation in the context of catabolic stress associated with non-specific illness.

Substrate-enzyme competition attenuates upregulated anaplerotic flux through malic enzyme in hypertrophied rat heart and restores triacylglyceride content: attenuating upregulated anaplerosis in hypertrophy.

Recent work identifies the recruitment of alternate routes for carbohydrate oxidation, other than pyruvate dehydrogenase (PDH), in hypertrophied heart. Increased carboxylation of pyruvate via cytosolic malic enzyme (ME), producing malate, enables "anaplerotic" influx of carbon into the citric acid cycle. In addition to inefficient NADH production from pyruvate fueling this anaplerosis, ME also consumes NADPH necessary for lipogenesis.

Effect of IL-1beta on survival and energy metabolism of R28 and RGC-5 retinal neurons.

Purpose: Interleukin-(IL)1beta expression is increased in the retina during a variety of diseases involving the death of retinal neurons and contributes to neurodegenerative processes through an unknown mechanism. This study was conducted to examine the effects of IL-1beta on the metabolism and viability of RGC-5 and R28 retinal neuronal cells.

Methods: Cellular reductive capacity was evaluated using WST-1 tetrazolium salt.

Mechanism of age-dependent susceptibility and novel treatment strategy in glutaric acidemia type I.

Glutaric acidemia type I (GA-I) is an inherited disorder of lysine and tryptophan metabolism presenting with striatal lesions anatomically and symptomatically similar to Huntington disease. Affected children commonly suffer acute brain injury in the context of a catabolic state associated with nonspecific illness. The mechanisms underlying injury and age-dependent susceptibility have been unknown, and lack of a diagnostic marker heralding brain injury has impeded intervention efforts.

Mitochondrial transport proteins of the brain.

In this study, cellular distribution and activity of glutamate and gamma-aminobutyric acid (GABA) transport as well as oxoglutarate transport across brain mitochondrial membranes were investigated. A goal was to establish cell-type-specific expression of key transporters and enzymes involved in neurotransmitter metabolism in order to estimate neurotransmitter and metabolite traffic between neurons and astrocytes. Two methods were used to isolate brain mitochondria.

Hyperglycemia, maturity-onset obesity, and insulin resistance in NONcNZO10/LtJ males, a new mouse model of type 2 diabetes.

As a new mouse model of obesity-induced diabetes generated by combining quantitative trait loci from New Zealand Obese (NZO/HlLt) and Nonobese Nondiabetic (NON/LtJ) mice, NONcNZO10/LtJ (RCS10) male mice developed type 2 diabetes characterized by maturity onset obesity, hyperglycemia, and insulin resistance. To metabolically profile the progression to diabetes in preobese and obese states, a 2-h hyperinsulinemic euglycemic clamp was performed and organ-specific changes in insulin action were assessed in awake RCS10 and NON/LtJ (control) males at 8 and 13 wk of age. Prior to development of obesity and attendant increases in hepatic lipid content, 8-wk-old RCS10 mice developed insulin resistance in liver and skeletal muscle due to significant decreases in insulin-stimulated glucose uptake and GLUT4 expression in muscle.

Recruitment of compensatory pathways to sustain oxidative flux with reduced carnitine palmitoyltransferase I activity characterizes inefficiency in energy metabolism in hypertrophied hearts.

Background: Transport rates of long-chain free fatty acids into mitochondria via carnitine palmitoyltransferase I relative to overall oxidative rates in hypertrophied hearts remain poorly understood. Furthermore, the extent of glucose oxidation, despite increased glycolysis in hypertrophy, remains controversial. The present study explores potential compensatory mechanisms to sustain tricarboxylic acid cycle flux that resolve the apparent discrepancy of reduced fatty acid oxidation without increased glucose oxidation through pyruvate dehydrogenase complex in the energy-poor, hypertrophied heart.

Energy sources for glutamate neurotransmission in the retina: absence of the aspartate/glutamate carrier produces reliance on glycolysis in glia.

The mitochondrial transporter, the aspartate/glutamate carrier (AGC), is a necessary component of the malate/aspartate cycle, which promotes the transfer into mitochondria of reducing equivalents generated in the cytosol during glycolysis. Without transfer of cytosolic reducing equivalents into mitochondria, neither glucose nor lactate can be completely oxidized. In the present study, immunohistochemistry was used to demonstrate the absence of AGC from retinal glia (Müller cells), but its presence in neurons and photoreceptor cells.

Diabetic retinopathy: seeing beyond glucose-induced microvascular disease.

Diabetic retinopathy remains a frightening prospect to patients and frustrates physicians. Destruction of damaged retina by photocoagulation remains the primary treatment nearly 50 years after its introduction. The diabetes pandemic requires new approaches to understand the pathophysiology and improve the detection, prevention, and treatment of retinopathy.

Analysis of glucose metabolism in diabetic rat retinas.

This study was conceived in an effort to understand cause and effect relationships between hyperglycemia and diabetic retinopathy. Numerous studies show that hyperglycemia leads to oxidative stress in the diabetic retinas, but the mechanisms that generate oxidative stress have not been resolved. Increased electron pressure on the mitochondrial electron transfer chain, increased generation of cytosolic NADH, and decreases in cellular NADPH have all been cited as possible sources of reactive oxygen species and nitrous oxide.

Branched-chain [corrected] amino acid metabolism: implications for establishing safe intakes.

There are several features of the metabolism of the indispensable BCAAs that set them apart from other indispensable amino acids. BCAA catabolism involves 2 initial enzymatic steps that are common to all 3 BCAAs; therefore, the dietary intake of an individual BCAA impacts on the catabolism of all 3. The first step is reversible transamination followed by irreversible oxidative decarboxylation of the branched-chain alpha-keto acid transamination products, the branched chain alpha-keto acids (BCKAs).

Minocycline reduces proinflammatory cytokine expression, microglial activation, and caspase-3 activation in a rodent model of diabetic retinopathy.

Diabetes leads to vascular leakage, glial dysfunction, and neuronal apoptosis within the retina. The goal of the studies reported here was to determine the role that retinal microglial cells play in diabetic retinopathy and assess whether minocycline can decrease microglial activation and alleviate retinal complications. Immunohistochemical analyses showed that retinal microglia are activated early in diabetes.

Neuroglial metabolism in the awake rat brain: CO2 fixation increases with brain activity.

Glial cells are thought to supply energy for neurotransmission by increasing nonoxidative glycolysis; however, oxidative metabolism in glia may also contribute to increased brain activity. To study glial contribution to cerebral energy metabolism in the unanesthetized state, we measured neuronal and glial metabolic fluxes in the awake rat brain by using a double isotopic-labeling technique and a two-compartment mathematical model of neurotransmitter metabolism. Rats (n = 23) were infused simultaneously with 14C-bicarbonate and [1-13C]glucose for up to 1 hr.

Evaluation of brain mitochondrial glutamate and alpha-ketoglutarate transport under physiologic conditions.

Some models of brain energy metabolism used to interpret in vivo (13)C nuclear magnetic resonance spectroscopic data assume that intramitochondrial alpha-ketoglutarate is in rapid isotopic equilibrium with total brain glutamate, most of which is cytosolic. If so, the kinetics of changes in (13)C-glutamate can be used to predict citric acid cycle flux. For this to be a valid assumption, the brain mitochondrial transporters of glutamate and alpha-ketoglutarate must operate under physiologic conditions at rates much faster than that of the citric acid cycle.

Whole-brain glutamate metabolism evaluated by steady-state kinetics using a double-isotope procedure: effects of gabapentin.

Cerebral rates of anaplerosis are known to be significant, yet the rates measured in vivo have been debated. In order to track glutamate metabolism in brain glutamatergic neurons and brain glia, for the first time unrestrained awake rats were continuously infused with a combination of H14CO3- and [1 - 13C]glucose in over 50 infusions ranging from 5 to 60 min. In whole-brain extracts from these animals, the appearance of 14C in brain glutamate and glutamine and appearance of 13C in the C-4 position of glutamate and glutamine were measured as a function of time.

Limited transfer of cytosolic NADH into mitochondria at high cardiac workload.

Glycolysis supplements energy synthesis at high cardiac workloads, producing not only ATP but also cytosolic NADH and pyruvate for oxidative ATP synthesis. Despite adequate Po(2), speculation exists that not all cytosolic NADH is oxidized by the mitochondria, leading to lactate production. In this study, we elucidate the mechanism for limited cytosolic NADH oxidation and increased lactate production at high workload despite adequate myocardial blood flow and oxygenation.