High energy phosphate compounds and ATPase activity of mitochondria in the brain of rats of different ages

Adenosine phosphate and creatine phosphate amount was determined in the brain tissue of 3-4-week, 6-8 month and 26-26 month old mongrel female rats. The maximum ATP and creatine phosphate amount and the minimum of ADP and AMP were found in young rats. In adult rats as compared with the young the ATP amount is the same, the ADP and AMP level rises, that of creative phosphate falls and energy charge decreases. In the brain of old rats the ATP and ADP amount falls, that of creatine phosphate and AMP remains at the level of mature-age animals. Despite a decrease in the ATP amount in the brain at the old age, the Mg, DNP- ATPase activity of mitochondria isolated from brain cortex and stem of the old rats remains at the level typical of adult animals.     

Metabolic Changes in Cerebral Cortex, Hippocampus, and Cerebellum During Sustained Bicuculline-Induced Seizures

Abstract— The objective of the present experiments was to study metabolic correlates to the localization of neuronal lesions during sustained seizures. To that end, status epilepticus was induced by i.v. administration of bicuculline in immobilized and artificially ventilated rats, since this model is known to cause neuronal cell damage in cerebral cortex and hippocampus but not in the cerebellum. After 20 or 120 min of continuous seizure activity, brain tissue was frozen in situ through the skull bone, and samples of cerebral cortex, hippocampus, and cerebellum were collected for analysis of glycolytic metabolites, phosphocreatine (PCr), ATP, ADP, AMP, and cyclic nucleotides. After 20 min of seizure activity, the two “vulnerable” structures (cerebral cortex and hippocampus) and the “resistant” one (cerebellum) showed similar changes in cerebral metabolic state, characterized by decreased tissue concentrations of PCr, ATP, and glycogen, and increased lactate concentrations and lactate/ pyruvate ratios. In all structures, though, the adenylate energy charge remained close to control. At the end of a 2-h period of status epilepticus, a clear deterioration of the energy state was observed in the cerebral cortex and the hippocampus, but not in the cerebellum. The reduction in adenylate energy charge in the cortex and hippocampus was associated with a seemingly paradoxical decrease in tissue lactate levels and with failure of glycogen resynthesis (cerebral cortex). Experiments with infusion of glucose during the second hour of a 2-h period of status epilepticus verified that the deterioration of tissue energy state was partly due to reduced substrate supply; however, even in animals with adequate tissue glucose concentrations, the energy charge of the two structures was significantly lowered. The cyclic nucleotides (cAMP and cGMP) behaved differently. Thus, whereas cAMP concentrations were either close to control (hippocampus and cerebellum) or moderately increased (cerebral cortex), the cGMP concentrations remained markedly elevated throughout the seizure period, the largest change being observed in the cerebellum. It is concluded that although the localization of neuronal damage and perturbation of cerebral energy state seem to correlate, the results cannot be taken as. evidence that cellular energy failure is the cause of the damage. Thus, it appears equally probable that the pathologically enhanced neuronal activity (and metabolic rate) underlies both the cell damage and the perturbed metabolic state. The observed changes in cyclic nucleotides do not appear to bear a causal relationship to the mechanisms of damage.     

Role of the biological activity of serotonin in the production of the "shock lung" syndrome]

The influence of the serotin biological activity on forming the "shock" lung syndrome was revealed in experiments on rats. Tachyhyperpnea with predominance of functional emphysema and a small number of atelectic tissue areas were observed in the animals with traumatic "shock" during the serotonin hypersecretion. Tachyhypopnea with a significant predominance of atelectiv areas was seen during the serotonin hypoproduction.     

The Influence of Hypoxia on the Concentrations of Cyclic Nucleotides in the Rat Brain

Abstract: In order to study the influence of hypoxia on cyclic nucleotides in the brain, we reduced arterial Po, for 15–30 min in lightly anaesthetised and artificially ventilated rats to obtain values ranging from about 45 to about 10 mm Hg. In an additional group (arterial Po₂ 18–22 mm Hg), the tissue hypoxia was aggravated by moderate arterial hypotension (mean arterial blood pressure about 80 mm Hg). In all animals, electrocortical activity was recorded. Cyclic GMP concentrations in cerebral cortex were unchanged in all groups but one. In that group, in which tissue hypoxia was severe enough to induce a suppression-burst EEG pattern and a measurable reduction in the adenylate energy charge, cyclic GMP concentrations were slightly increased ( p < 0.05). Cyclic AMP concentrations remained unaltered at all degrees of hypoxia studied. It is concluded that changes in cyclic nucleotides in brain tissue occur first at such severe degrees of hypoxia of the duration studied that function and metabolism are profoundly altered.     

CYCLIC NUCLEOTIDES IN MURINE BRAIN: EFFECT OF HYPOTHERMIA ON ADENOSINE 3'',5'' MONOPHOSPHATE, GLYCOGEN PHOSPHORYLASE, GLYCOGEN SYNTHASE AND METABOLITES FOLLOWING MAXIMAL ELECTROSHOCK OR DECAPITATION

Abstract —The accumulation of adenosine-3',5'-cyclic monophosphate (cyclic AMP) has been investigated in murine brain following electroconvulsive shock and decapitation. Animals were made hypothermic (20°C) to minimize the freezing time of the brain and to delay metabolic events. Cyclic AMP concentrations were decreased in the cerebral cortex of hypothermic rats and mice. Furthermore, the changes in cyclic AMP elicited by electroconvulsive shock and decapitation were delayed. In hypothermic animals, the metabolic rate as determined by high energy phosphate use was decreased to 65% of control values. The interconversions of the active and inactive forms of glycogen phosphorylase and glycogen synthase were sufficiently retarded in hypothermic animals to correlate with changes in cyclic AMP concentrations. The conversion of phosphorylase b to a and synthase a to b occurred when cyclic AMP concentrations had increased from 2 to 5 μmol/kg, following either electroconvulsive shock or decapitation. The results indicate that cyclic AMP plays a role in regulation of glycogen metabolism in cerebral cortex.     

Regional Levels of Lactate and Norepinephrine After Experimental Brain Injury

Abstract: The recently developed controlled cortical impact model of brain injury in rats may be an excellent tool by which to attempt to understand the neurochemical mechanisms mediating the pathophysiology of traumatic brain injury. In this study, rats were subjected to lateral controlled cortical impact brain injury of low grade severity; their brains were frozen in situ at various times after injury to measure regional levels of lactate, high energy phosphates, and norepinephrine. Tissue lactate concentration in the injury site left cortex was increased in injured animals by sixfold at 30 min and twofold at 2.5 h and 24 h after injury ( p < 0.05). At all postinjury times, lactate concentration was also increased in injured animals by about twofold in the cortex and hippocampus adjacent to the injury site ( p < 0.05). No significant changes occurred in the levels of ATP and phosphocreatine in most of the brain regions of injured animals. However, in the primary site of injury (left cortex), phosphocreatine concentration was decreased by 40% in injured animals at 30 min after injury ( p < 0.05). The norepinephrine concentration was decreased in the injury site left cortex of injured animals by 38% at 30 min, 29% at 2.5 h, and 30% at 24 h after injury ( p < 0.05). The level of norepinephrine was also reduced by ∼20% in the cortex adjacent to the injury site in injured animals. The present results suggest that controlled cortical impact brain injury produces disorder in the neuronal oxidative and norepinephrine metabolism.     

Phosphorylation of Myristoylated Alanine-Rich C Kinase Substrate (MARCKS) by Proline-Directed Protein Kinases and Its Dephosphorylation

Abstract: Excitatory amino acid (EAA) neurotransmitters may play a role in the pathophysiology of traumatic injury to the CNS. Although NMDA receptor antagonists have been reported to have therapeutic efficacy in animal models of brain injury, these compounds may have unacceptable toxicity for clinical use. One alternative approach is to inhibit the release of EAAs following traumatic injury. The present study examined the effects of administration of a novel sodium channel blocker and EAA release inhibitor, BW1003C87, or the NMDA receptor-associated ion channel blocker magnesium chloride on cerebral edema formation following experimental brain injury in the rat. Animals (n = 33) were subjected to fluid percussion brain injury of moderate severity (2.3 atm) over the left parietal cortex. Fifteen minutes after injury, the animals received a constant infusion of BW1003C87 (10 mg/kg, i.v.), magnesium chloride (300 µmol/kg, i.v.), or saline over 15 min (2.75 ml/kg/15 min). In all animals, regional tissue water content in brain was assessed at 48 h after injury, using the wet weight/dry weight technique. In saline-treated control animals, fluid percussion brain injury produced significant regional brain edema in injured left parietal cortex ( p < 0.001), the cortical area adjacent to the site of maximal injury ( p < 0.001), left hippocampus ( p < 0.001), and left thalamus ( p = 0.02) at 48 h after brain injury. Administration of BW1003C87 15 min postinjury significantly reduced focal brain edema in the cortical area adjacent to the site of maximal injury ( p < 0.02) and left hippocampus ( p < 0.01), whereas magnesium chloride attenuated edema in left hippocampus ( p = 0.02). These results suggest that excitatory neurotransmission may play an important role in the pathogenesis of posttraumatic brain edema and that pre- or post-synaptic blockade of glutamate receptor systems may attenuate part of the deleterious sequelae of traumatic brain injury.     

Electric shock convulsions in the rabbit and brain cortex GABAA receptor function

The effect of electric shock convulsions (ESC) on the function of brain cortex GABA_A receptors has been studied in the rabbit. Three single electroconvulsive shocks (ECS) were given at intervals of 48 hours and the brain cortex was sampled 36 hours after the last shock. The dose-response curve was determined for GABA-stimulated³⁶Cl^– accumulation into brain cortex microsacs. The parameters of the curve (maximal accumulation rate, K_a and Hill coefficient, n) were constant when determined in two different series of experiences. Animals handled in the same way as the animals from the electric shock group but which did not receive the ECSs (sham ECS group) showed similar maximal accumulation rate and K_a. However, theaverage n coefficient was significantly higher in the electric shock group. Naive animals, taken from their cages just before the sacrifice, showed dose-response curves which varied from one experimental series to another. This last result (confirming previous observations) shows modifications and inconsistencies in the evaluation of GABA_A receptor function in stressed handling-naive animals.Clinica Neurologica dell'Universita.     

Mechanics of blast loading on the head models in the study of traumatic brain injury using experimental and computational approaches

Blast waves generated by improvised explosive devices can cause mild, moderate to severe traumatic brain injury in soldiers and civilians. To understand the interactions of blast waves on the head and brain and to identify the mechanisms of injury, compression-driven air shock tubes are extensively used in laboratory settings to simulate the field conditions. The overall goal of this effort is to understand the mechanics of blast wave–head interactions as the blast wave traverses the head/brain continuum. Toward this goal, surrogate head model is subjected to well-controlled blast wave profile in the shock tube environment, and the results are analyzed using combined experimental and numerical approaches. The validated numerical models are then used to investigate the spatiotemporal distribution of stresses and pressure in the human skull and brain. By detailing the results from a series of careful experiments and numerical simulations, this paper demonstrates that: (1) Geometry of the head governs the flow dynamics around the head which in turn determines the net mechanical load on the head. (2) Biomechanical loading of the brain is governed by direct wave transmission, structural deformations, and wave reflections from tissue–material interfaces. (3) Deformation and stress analysis of the skull and brain show that skull flexure and tissue cavitation are possible mechanisms of blast-induced traumatic brain injury.     

Cutaneous shock produces correlated shifts in slow potential amplitude and cyclic 3',5'-adenosine monophosphate level in the parietal cortex of the conscious rat.

Abstract— Fourteen animals each received 4 cutaneous shocks with an interval of 3–5 min between them. During a fifth trial 3–5 min later, eleven subjects received a fifth shock and then 3–30 s afterwards, as cerebral slow potentials developed in response to the stimulus, samples of parietal cortex were rapidly frozen and extracted by a cryoplate. Three baseline subjects received no shock at the time of the fifth trial and had their parietal tissue samples taken without the presence of slow potentials. A correlation coefficient of r=?0.77 (P < 0.01) was observed between the slow potential amplitude on the surface of the parietal cortex at the time of the sampling and the analyzed level of cyclic AMP in the underlying tissue. Five of the shocked animals whose samples were taken before the slow potentials increased significantly showed a tissue level of 11.1 ± 3.0 pmol cyclic AMP/mg protein. This level was significantly higher (P < 0.01) than that of the baseline animals (3.1 ± 2.0 pmol cyclic AMP/mg protein). The other six shocked animals who had developed large slow potentials manifested a cyclic AMP level that was not different from the baseline group. It is concluded that a reoccurring cutaneous shock results in the immediate increase in the level of cyclic AMP in the parietal cortex and that within 30 s this level decreases in proportion to the amplitude of the slow potential that develops in the same region.     

Intracellular oxygen tension and energy metabolism in the cat brain cortex during haemorrhagic shock.

The changes in intracellular oxygen tension and energy metabolism of the cat brain cortex were studied by surface fluororeflectometry during haemorrhagic shock. The results may be summarized as follows. (a) Intracellular oxygen tension, i.e. the maximum cortical NAD reduction obtained during nitrogen gas inhalation decreased gradually during the hypovolaemic phase of shock and finally, the brain cortex became ischaemic. (b) Partial uncoupling of the cerebrocortical mitochondrial respiration and oxidative phosphorylation appeared in the very early period of bleeding, as indicated by the overshot of the cortical NAD/NADH redox state towards oxidation subsequent to the cessation of nitrogen gas inhalation. Partial uncoupling of mitochondrial respiration and oxidative phosphorylation became more pronounced during the later phases of bleeding, finally, the mitochondrial electron transport stopped. In line with these changes the frequency and the amplitude of ECoG decreased gradually and markedly during the hypovolaemic phase of shock. (c) Microcirculation and energy metabolism of the cat brain cortex were severely and irreversibly damaged during the hypovolaemic phase of shock. This was clearly shown by the fact that in the majority of experiments the nitrogen anoxia after reinfusion failed to bring about changes in the cortical NAD/NADH redox state and the ECoG changes occurred during bleeding did not improve after reinfusion. It is concluded that the early disturbances of cerebrocortical energy metabolism play an important role in the development of neural and vascular lesions of the brain that occur during haemorrhagic shock.     

Effects of maternal thiamine deficiency on the development of thiamine-dependent enzymes in regions of the rat brain

Previous studies suggest that developing rat brain is susceptible to reduced thiamine intake. In order to assess the metabolic basis for this susceptibility, activities of three thiamine-dependent enzymes (pyruvate dehydrogenase complex, -ketoglutarate dehydrogenase and transketolase) were measured in homogenates of brain tissue from the offspring of thiamine-deficient mothers. Control groups of animals were pair-fed to equal food consumption with the thiamine-deficient animals. The study revealed region-selective delays in the establishment of adult activities of thiamine-dependent enzymes as a result of maternal thiamine deficiency. Pyruvate dehydrogenase complex activities in cerebral cortex were significantly reduced (by 20% P < 0.05); -ketoglutarate dehydrogenase activities were also reduced in cerebral cortex (by 30% P < 0.05). In the case of transketolase, enzyme activities were significantly reduced in cerebral cortex, cerebellum and brainstem. Following thiamine replenishment, defective enzyme activities were restored to normal in all cases. However, since thiamine-dependent enzymes are important for the establishment of adult patterns of cerebral energy metabolism and also in myelin synthesis, maternal thiamine deficiency resulting in reductions of thiamine-dependent enzymes at a vulnerable period in brain development could have serious metabolic consequences leading to permanent neurological sequellae in the offspring.     

Oxidative stress following traumatic brain injury in rats: quantitation of biomarkers and detection of free radical intermediates

Oxidative stress may contribute to many pathophysiologic changes that occur after traumatic brain injury. In the current study, contemporary methods of detecting oxidative stress were used in a rodent model of traumatic brain injury. The level of the stable product derived from peroxidation of arachidonyl residues in phospholipids, 8-epi-prostaglandin F(2alpha), was increased at 6 and 24 h after traumatic brain injury. Furthermore, relative amounts of fluorescent end products of lipid peroxidation in brain extracts were increased at 6 and 24 h after trauma compared with sham-operated controls. The total antioxidant reserves of brain homogenates and water-soluble antioxidant reserves as well as tissue concentrations of ascorbate, GSH, and protein sulfhydryls were reduced after traumatic brain injury. A selective inhibitor of cyclooxygenase-2, SC 58125, prevented depletion of ascorbate and thiols, the two major water-soluble antioxidants in traumatized brain. Electron paramagnetic resonance (EPR) spectroscopy of rat cortex homogenates failed to detect any radical adducts with a spin trap, 5,5-dimethyl-1-pyrroline N:-oxide, but did detect ascorbate radical signals. The ascorbate radical EPR signals increased in brain homogenates derived from traumatized brain samples compared with sham-operated controls. These results along with detailed model experiments in vitro indicate that ascorbate is a major antioxidant in brain and that the EPR assay of ascorbate radicals may be used to monitor production of free radicals in brain tissue after traumatic brain injury.     

Morpho-functional study of the brain of animals with different types of individual resistance to hypoxia

The experiments on white rats have shown that animals with distinct tolerance to hypoxia were characterized by individual metabolic changes in phylogenetically different brain structures. Adaptation to hypoxia in animals with high tolerance was associated with metabolic changes in the reticular formation and in animals with low tolerance with changes in the cerebral cortex. The experiments have shown that white rats with distinct individual tolerance to hypoxia are characterized by an inherent level of plastic metabolism in different brain structures. A correlation between brain tissue metabolism and individual tolerance of animals to hypoxia is suggested.     

Creatine-Enhanced Diet Alters Levels of Lactate and Free Fatty Acids After Experimental Brain Injury

Free fatty acids (FFA) and lactic acid are markers of secondary cellular injury following traumatic brain injury (TBI). We previously showed that animals fed a creatine (Cr)-enriched diet are afforded neuroprotection following TBI. To further characterize the neuroprotective Cr diet, we studied neurochemical changes in cortex and hippocampus following a moderate injury. Adult rats were fed either a control or Cr-supplemented diet (0.5%, 1%) for 2 weeks before TBI. At 30 min or 6 h after injury, tissue was processed for quantitative analysis of neurochemical changes. Both lactate and FFA were significantly increased in all tissues ipsilateral to the injury. Cr-fed animals had significantly lower levels, although the levels were elevated compared to sham controls. Animals fed a 1% Cr-diet were afforded greater neuroprotection than animals fed a 0.5% Cr diet. These results support the idea that a Cr-enriched diet can provide substantial neuroprotection in part by suppressing secondary brain injury.     

Manganese and ammonia interactions in the brain of cirrhotic rats: effects on brain ammonia metabolism

Hepatic encephalopathy is a major complication of cirrhosis. Ammonia and manganese have been associated with hepatic encephalopathy underlying mechanisms. Motor impairment and brain edema are common signs of hepatic encephalopathy. In the present study a model of liver damage in rats was combined with ammonia and manganese exposure to evaluate the role of these substances separately and their interactions on brain glutamine, water content and motor coordination. Additionally, we explored brain levels of each substance -Mn and ammonia- in the presence or absence of the other. Liver damage was induced by bile duct ligation. Rats were exposed to MnCl₂ in drinking water (1 mg Mn/ml) and to ammonia in chow pellets containing 20% ammonium acetate (w/w). As expected, manganese and ammonia levels increased in the brain of cirrhotic rats exposed to these substances; in these animals, glutamine brain levels also increased and positively correlated with tissue water content in cortex. A three way-ANOVA showed that manganese favored ammonia and glutamine accumulation in brain, and possibly their subsequent deleterious effects, as evidenced by the fact that manganese and ammonia accumulation in the brain of cirrhotic rats severely affected motor function. These results suggest that even when controlling ammonia levels in cirrhotic patients, reduction of manganese intake is also a potential strategy to be considered in clinical practice.     

Epidermal growth factor receptor ontogeny in mice with congenital hypothyroidism

To assess the effect of endogenous thyroid hormone on hepatic EGF receptors in developing mice we measured EGF binding to plasma membrane receptors in liver and brain of mice with congenital hypothyroidism and in euthyroid controls at 20, 30 and 40 days of age. At 20 days hepatic EGF receptor binding was low in both hypothyroid and control animals. Between 20 and 30 days the hepatic binding increased dramatically in the euthyroid animals, an increase that was greater in males than females. The increase in binding was due to an increase in the high affinity receptor population. Among hypothyroid animals there were no changes in hepatic EGF receptor binding with increasing age. In cerebral cortex EGF binding was similar in euthyroid and hypothyroid animals at all ages. These results suggest that thyroxine has regulatory effects on the postnatal ontogeny of hepatic EGF receptors.     

Lactography as an approach to monitor glucose metabolism on-line in brain and muscle

1. Thus far metabolic processes in the intact animal (or man) have been studied either by the analysis of body fluids, of biopsies, of tissue obtained post mortem or by techniques, requiring dedicated and expensive equipment (such as positron emission tomography or magnetic resonance spectroscopy). 2. Here we describe a relatively simple and inexpensive technique, that can be applied in vivo to study metabolism in brain regions and muscle in the freely moving rat and in human peripheral tissue. 3. The method is based on microdialysis allowing continuous sampling from the extracellular space, the enzymatic conversion of lactate and the on-line detection of fluorescent NADH. 4. Examples of the application of our technique include the monitoring of lactate efflux from various brain regions of behaving animals under a variety of stress exposures, during ischemia or hypoxia and drug treatments. 5. The results indicate that in brain lactate is not exclusively formed under hypoxia and that neuronal activation leads also to lactate formation, possibly due to the compartmentation of both the involved enzymes and the energy metabolism. 6. The increase of lactate formation in contracting or ischemic muscle or during exercise could also be followed on-line in the rat, suggesting that our approach allows the continuous monitoring of anaerobic metabolism in man e.g. during traumatic or arteriosclerotic limb ischemia or lactic acidosis in shock states. 7. The principle of our approach can easily be adapted to other metabolites, thus enabling to monitor other metabolic pathways in vivo as well.     

Reactivity of the cerebrocortical vasculature and energy metabolism to direct cortical stimulation in haemorrhagic shock.

Direct electrical stimulation of the cerebral cortex was used to determine the changes of cortical carbohydrate and oxidative metabolism and of vascular reactivity during haemorrhagic shock. The results were as follows. 1. Electrical stimulation of the brain cortex applied in the control period led to a marked vasodilation and NAD reduction that was preceded in part of the experiments by a transient NADH oxidation. It is suggested that the increase in cortical NADH fluorescence observed during direct stimulation is due to the fact that the rate of cytoplasmic NADH production exceeded the rate of mitochondrial NADH oxidation and of the rate of H+-transport from the cytoplasm into the mitochondria. 2. The cerebrocortical vascular and NAD/NADH redox state responses induced by electrical stimulation changed in the early hypovolaemic phase of shock. At this time, electrical stimulation of the brain cortex led to NADH oxidation in the majority of the experiments or in some experiments, the stimulation did not bring about changes in the redox state of the cortex. The total loss of the reactivity to direct stimulation of the cerebrocortical vessels and of energy metabolism preceded the occurrence of cortical ischaemia during the hypovolaemic phase of shock. 3. Since after reinfusion of the shed blood, redox state and vasculature remained unresponsive to stimulation even in those experiments in which the cortical ischaemia improved, it is concluded that the carbohydrate and oxidative metabolism of the brain cortex were already irreversibly damaged in the early phase of hypovolaemic shock.     

L-Aspartate,L-Ornithine and L-Ornithine-L-Aspartate (LOLA) and Their Impact on Brain Energy Metabolism

L-Ornithine-L-aspartate (LOLA), a crystalline salt, is used primarily in the management of hepatic encephalopathy. The degree to which it might penetrate the brain, and the effects it might have on metabolism in brain are poorly understood. Here, to investigate the effects of LOLA on brain energy metabolism we incubated brain cortical tissue slices from guinea pig (Cavea porcellus) with the constituent amino acids of LOLA, L-ornithine or L-aspartate, as well as LOLA, in the presence of [1-¹³C]D-glucose and [1,2-¹³C]acetate; these labelled substrates are useful indicators of brain metabolic activity. L-Ornithine produced significant “sedative” effects on brain slice metabolism, most likely via conversion of ornithine to GABA via the ornithine aminotransferase pathway, while L-aspartate showed concentration-dependent excitatory effects. The metabolic effects of LOLA reflected a mix of these two different processes and were concentration-dependent. We also investigated the effect of an intraperitoneal bolus injection of L-ornithine, L-aspartate or LOLA on levels of metabolites in kidney, liver and brain cortex and brain stem in mice (C57Bl6J) 1 h later. No significant changes in metabolite levels were seen following the bolus injection of L-aspartate, most likely due to rapid metabolism of aspartate before reaching the target tissue. Brain cortex glutamate was decreased by L-ornithine but no other brain effects were observed with any other compound. Kidney levels of aspartate were increased after injection of L-ornithine and LOLA which may be due to interference by ornithine with the kidney urea cycle. It is likely that without optimising chronic intravenous infusion, LOLA has minimal impact on healthy brain energy metabolism due to systemic clearance and the blood - brain barrier.