Asunto(s)

Asunto(s)

RESUMEN

The rates of uptake and oxidation of [U-(14)C]lactate and [U-(14)C]glucose were determined in primary cultures of astrocytes and neurons from rat brain, in the presence and absence of the monocarboxylic acid transport inhibitor alpha-cyano-4-hydroxycinnamate (4-CIN). The rates of uptake for 1 mM lactate and glucose were 7.45 +/- 1.35 and 8.80 +/- 1.0 nmol/30 sec/mg protein in astrocytes and 2.36 +/- 0.19 and 1.93 +/- 0.16 nmol/30 sec/mg protein in neuron cultures, respectively. Lactate transport into both astrocytes and neurons was significantly decreased by 0.25-1.0 mM 4-CIN; however, glucose uptake was not affected. The rates of (14)CO(2) formation from 1 mM lactate and glucose were 12.49 +/- 0.77 and 3.42 +/- 0.67 nmol/hr/mg protein in astrocytes and 29.32 +/- 2.81 and 10.04 +/- 1.79 nmol/hr/mg protein in neurons, respectively. Incubation with 0.25 mM 4-CIN decreased the oxidation of lactate and glucose to 57.1% and 54.1% of control values in astrocytes and to 13.2% and 41.6% of the control rates in neurons, respectively. Preincubation with 4-CIN further decreased the oxidation of both glucose and lactate. Studies with glucose specifically labeled in the one and six positions demonstrated that 4-CIN decreased mitochondrial glucose oxidation but did not impair the metabolism of glucose via the pentose phosphate pathway in the cytosol. The lack of effect of 4-CIN on glutamate oxidation demonstrated that overall mitochondrial metabolism was not impaired. These findings suggest that the impaired neuronal function and tissue damage in the presence of 4-CIN observed in other studies may be due in part to decreased uptake of lactate; however, the effects of 4-CIN on mitochondrial transport would significantly decrease the oxidative metabolism of pyruvate derived from both glucose and lactate.

Asunto(s)

RESUMEN

To determine lung malic enzyme activity at varying stages of development, both cytosolic and mitochondrial enzyme activities were assayed in rat lungs at various stages from day 16 of fetal life to 2 months of postnatal life by measuring the production of 14CO2 from 14C-malate. Malic enzyme activities were significantly higher in the mitochondrial than in the cytosolic fractions at all ages studied. The mitochondrial malic enzyme activity was significantly higher in canalicular stage (days 19-20) stage of lung development when compared to the glandular stage (days 16-18). The mitochondrial fraction at day 19 exhibited biphasic kinetics: high affinity, Km = 0.45 mmol, Vmax = 10.04 nmol/mg protein/min; and low affinity, Km = 5.48 mmol, Vmax = 56.83 nmol/mg protein/min. The cytosolic malic enzyme activity of all fetal stages (saccular stage [days 16-18], canalicular stage [days 19-20], and glandular stage [days 21-22] were significantly higher when compared to postnatal levels (postnatal days 1-10, adult). In contrast to the mitochondrial fraction, at day 19, the cytosolic fraction showed a single Km of 0.23 mmol, Vmax = 12.32 nmol/mg protein/min. The increased mitochondrial malic enzyme activity during late gestation would suggest that, as we have previously demonstrated, anaplerotic substrates other than glucose, may provide a significant energy source in fetal lung. The increased cytosolic activity in the prenatal phases would suggest that the NADPH provided from malic enzyme is an important contributor to de novo fatty acid synthesis, leading to surfactant synthesis, critical to normal lung development in late gestation.

Asunto(s)

Asunto(s)

RESUMEN

There have been numerous studies on the activity and localization of aspartate aminotransferase (AAT) and glutamate dehydrogenase (GDH) in brain tissue. However, there is still a controversy as to the specific roles and relative importance of these enzymes in glutamate and glutamine metabolism in astrocytes and neurons or synaptic terminals. There are many reports documenting GDH activity in synaptic terminals, yet the misconception that it is a glial enzyme persists. Furthermore, there is evidence that this tightly regulated enzyme may have an increased role in synaptic metabolism in adverse conditions such as low glucose and hyperammonemia that could compromise synaptic function. In the present study, we report high activity of both AAT and GDH in mitochondrial subfractions from cortical synaptic terminals. The relative amount of GDH/AAT activity was higher in SM2 mitochondria, compared to SM1 mitochondria. Such a differential distribution of enzymes can contribute significantly to the compartmentation of metabolism. There is evidence that the metabolic capabilities of the SM1 and SM2 subfractions of synaptic mitochondria are compatible with the compartments A and B of neuronal metabolism proposed by Waagepetersen et al. (1998b. Dev. Neurosci. 20, 310-320).

Asunto(s)

RESUMEN

Most of the malic enzyme activity in the brain is found in the mitochondria. This isozyme may have a key role in the pyruvate recycling pathway which utilizes dicarboxylic acids and substrates such as glutamine to provide pyruvate to maintain TCA cycle activity when glucose and lactate are low. In the present study we determined the activity and kinetics of malic enzyme in two subfractions of mitochondria isolated from cortical synaptic terminals, as well as the activity and kinetics in mitochondria isolated from primary cultures of cortical neurons and cerebellar granule cells. The synaptic mitochondrial fractions had very high mitochondrial malic enzyme (mME) activity with a Km and a Vmax of 0.37 mM and 32.6 nmol/min/mg protein and 0.29 mM and 22.4 nmol/min mg protein, for the SM2 and SM1 fractions, respectively. The Km and Vmax for malic enzyme activity in mitochondria isolated from cortical neurons was 0.10 mM and 1.4 nmol/min/mg protein and from cerebellar granule cells was 0.16 mM and 5.2 nmol/min/mg protein. These data show that mME activity is highly enriched in cortical synaptic mitochondria compared to mitochondria from cultured cortical neurons. The activity of mME in cerebellar granule cells is of the same magnitude as astrocyte mitochondria. The extremely high activity of mME in synaptic mitochondria is consistent with a role for mME in the pyruvate recycling pathway, and a function in maintaining the intramitochondrial reduced glutathione in synaptic terminals.

Asunto(s)

RESUMEN

Since lactate released by glial cells may be a key substrate for energy in neurons, the kinetics for the uptake of L-[U-14C]lactate by cortical synaptic terminals from 7- to 8-week-old rat brain were determined. Lactate uptake was temperature-dependent, and increased by 64.9% at pH 6.2, and decreased by 43.4% at pH 8.2 relative to uptake at pH 7.3. Uptake of monocarboxylic acids was saturable with increasing substrate concentration. Eadie-Hofstee plots of the data gave evidence of two carrier-mediated uptake mechanisms with a high-affinity Km of 0.66 mM and Vmax of 3.66 mM for pyruvate, and a low-affinity system with a Km of 9.9 mM for both lactate and pyruvate and Vmax values of 16.6 and 23.1 nmol/30 s/mg protein for lactate and pyruvate, respectively. Saturable uptake was seen in the presence of 10 mM alpha-cyano-4-hydroxycinnamate. Lactate transport by synaptic terminals was much more sensitive to inhibition by sulfhydryl reagents than transport in astrocytes. Addition of 0.5 and 2 mM mersalyl decreased the uptake of 1 mM lactate by synaptic terminals by 59.3 and 66.37%, respectively. Pyruvate moderately decreased lactate transport, whereas 3-hydroxybutyrate had little effect. Quercetin, an inhibitor of lactate release, had little effect on the content of 14C lactate in synaptic terminals, supporting the concept that the majority of lactate produced within brain is from glial cells. Oxidation of L-[U-14C]lactate by synaptosomes was saturable, and yielded a Km of 1.23 mM and a Vmax of 116 nmol/h/mg protein. Overall the studies show that synaptic terminals from adult brain have a high capacity for transport and oxidation of lactate, consistent with the proposed role for this compound in metabolic trafficking in brain. Furthermore, the data provide kinetic evidence of two carrier-mediated mechanisms for monocarboxylic acid transport by synaptosomes and demonstrate that uptake of lactate by synaptic terminals is regulated differently than transport by astrocytes. Uptake of lactate by synaptic terminals also has differences from the systems described for neurons.

Asunto(s)

RESUMEN

Paroxysmal tonic upgaze (PTU) of childhood is a distinctive neuro-ophthalmological syndrome of unknown etiology and pathogenesis that is characterized by episodes of sustained upward deviation of the eyes, often with incomplete downward saccades on attempted downgaze. It is generally regarded as having a benign outcome. We observed 16 children with PTU, from 10 months to 11 years from onset (mean, 5.4 years), to study the natural history and possible etiology. Five cases were from two unrelated families. Onset of PTU occurred either during or after an intercurrent infection or vaccination in 5 children. No antecedent was identifiable in the rest. PTU had completely resolved in 10 children (62%) (mean age at offset, 2.5 years), whereas 2 children intermittently manifest a modified form of the disorder. At follow-up, 11 children (69%) had developmental delay, intellectual disability, or language delay and 9 (56%) had ocular motility problems other than PTU. Only 3 children (19%) had normal development and neurological findings. PTU is a heterogeneous syndrome with respect to associations and outcome and may simply be an age-dependent manifestation of a variety of disorders affecting corticomesencephalic control of vertical eye movement. This disorder may be an early sign of more widespread neurological dysfunction.

Asunto(s)

RESUMEN

Sixty-three children with new-onset temporal lobe epilepsy (TLE) underwent extensive clinical, EEG, and neuroimaging investigation as part of a prospective, community-based cohort study of the natural history of TLE in childhood. Complex partial seizures occurred in 94% of the children, and tonic-clonic seizures occurred in 14%. Developmental, behavioral, or learning problems were present in 38%. Eighteen children (29%) had a significant illness/event prior to the onset of TLE, including febrile status epilepticus in seven, meningitis in four, respiratory arrest in two, and head injury in one. Magnetic resonance imaging or computed tomography revealed structural abnormalities of the temporal lobe in 24 children (38%), including hippocampal sclerosis (HS) in 13 and tumor in eight. There was a strong association between HS and a history of significant illness/event prior to the onset of TLE (p < 0.001). Analysis of past history and neuroimaging findings led us to propose three etiologically defined subgroups of TLE; developmental TLE (10 children with long-standing, nonprogressive temporal lobe tumors and malformations), TLE with HS/significant antecedents (18 children with HS or a history of a significant illness/event), and cryptogenic TLE (34 children with normal neuroimaging findings and no significant past history). Etiologic differences between children with new-onset TLE may confer prognostic information that will be useful for counselling families and planning treatment.

Asunto(s)

RESUMEN

Glucose has been thought to be the primary substrate for energy metabolism in the developing lung; however, alternate substrates are used for energy metabolism in other organs. To examine the role of alternate substrates in the lung, we measured rates of oxidation of glutamine, glucose, lactate, and 3-hydroxybutyrate in type II pneumocytes isolated from d 19 fetal rat lungs by measuring the production of 14CO2 from labeled substrates. Glutamine had a rate of 24.36 +/- 4.51 nmol 14CO2 produced/ h/mg of protein (mean +/- SEM), whereas lactate had a significantly higher rate, 40.29 +/- 4.42. 3-Hydroxybutyrate had a rate of 14.91 +/- 1.93. The rate of glucose oxidation was 2.13 +/- 0.36, significantly lower than that of glutamine. To examine the interactions of substrates normally found in the intracellular milieu, we measured the effect of unlabeled substrates as competitors on labeled substrate. This identifies multiple metabolic compartments of energy metabolism. Glucose, but not lactate, inhibited the oxidation of glutamine, suggesting a compartmentation of tricarboxylic acid cycle activity, rather than simple dilution by glucose. Glucose and lactate had reciprocal inhibition. Our data suggest at least two separate compartments in the type II cells for substrate oxidation, one for glutamine metabolism and a second for glucose metabolism. In summary, we have documented that glutamine and other alternate substrates are oxidized preferentially over glucose for energy metabolism in the d 19 fetal rat lung type II pneumocyte. In addition, we have delineated some of the compartmentation that occurs within the developing type II cell, which may determine how these substrates are used.

Asunto(s)

RESUMEN

Because multiple substrates have been shown to play a role in the metabolic homeostasis of different tissues, a series of studies were initiated to examine the role of alternate substrates in the lung. In these studies, we measured rates of oxidation of glutamine, glucose, lactate and 3-hydroxybutyrate in fibroblasts isolated from d 19 fetal rat lungs by measuring the production of 14CO2 from labeled substrates and compared them with earlier studies of isolated Type II cells. The rate of glutamine oxidation was 16.04 nmol 14CO2 x mg protein(-1) x hr(-1) in the fibroblasts compared with 24.36 in Type II cells. Three-hydroxybutyrate had a rate of 10.75 in the fibroblasts and 14.9 in the Type II cells. Lactate oxidation in fibroblasts was similar to that of glutamine, with a rate of 18.49; however, in Type II cells the rate of lactate oxidation was significantly higher at 40.29. Glucose was oxidized at a rate significantly lower than the other three substrates. In the fibroblasts, that rate was 1.22 and in Type II cells it was 2.13. To examine the interactions of substrates normally found in the intracellular milieu, we measured the effect of unlabeled substrates as competitors on labeled substrate in the fibroblasts, similar to our studies with Type II cells that identified multiple metabolic compartments of energy metabolism in these cell populations. Glucose, but not lactate, inhibited the oxidation of glutamine, suggesting a compartmentation of tricarboxylic acid cycle activity rather than simple dilution by glucose. Glucose and lactate had reciprocal inhibition in the Type II cells. Our data suggest at least two separate compartments in developing lung cells for substrate oxidation: one for glutamine metabolism and a second for glucose metabolism. In summary, we have documented that glutamine and other alternate substrates are oxidized preferentially over glucose for energy metabolism in the d 19 fetal rat lung.

Asunto(s)

Asunto(s)

Asunto(s)

RESUMEN

Idiopathic hypothalamic dysfunction is a rare but well-defined entity in childhood characterized by adipsia-hypernatremia, obesity, poor thermoregulation, and disturbance of pituitary function. Two cases of idiopathic hypothalamic dysfunction are described. There are 10 previously reported cases in the literature, and the clinical features are compared. The present cases are unique in that the patients also had bilaterally dilated unresponsive pupils. In the first case, there was no demonstrable pathology at autopsy; in the second case, lymphocytic infiltration of the hypothalamus and midbrain associated with neuronal loss was present at autopsy. Possible etiologies are discussed.

Asunto(s)

RESUMEN

It is well documented that the brain preferentially utilizes alternative substrates for energy during brain development; however, less is known about the use of these substrates by synaptic terminals. The present study compared the rates of 14CO2 production from 1 mM D-[6-14C]glucose, L-[U-14C]glutamine, D-3-hydroxy[3-14C]butyrate, L-[U-14C]lactate and L-[U-14C]malate by synaptic terminals isolated from 17- to 18-day-old and 7- to 8-week-old rat brain. The rates of 14CO2 production from glucose, glutamine, 3-hydroxybutyrate, lactate and malate were 8.55 +/- 0.78, 25.90 +/- 4.58, 42.28 +/- 3.54, 48.42 +/- 2.09, and 9.31 +/- 1.61 nmol/h/mg protein (mean +/- SEM), respectively, in synaptic terminals isolated from 17- to 18-day-old rat brain and 12.95 +/- 1.64, 30.62 +/- 4.19, 16.09 +/- 2.62, 40.33 +/- 6.77, and 8.25 +/- 1.69 nmol/h/mg protein (mean +/- SEM), respectively, in synaptic terminals isolated from 7- to 8-week-old rat brain. In competition studies using unlabelled added substrates, the addition of 3-hydroxybutyrate, lactate or glutamine greatly decreased the rate of 14CO2 production from labelled glucose. Added unlabelled glucose increased the rate of 14CO2 production from 3-hydroxybutyrate in synaptic terminals from 7- to 8-week-old rat brain, but had no effect on 14CO2 production from any other substrates. Lactate also increased 14CO2 production from 3-hydroxybutyrate at 7-8 weeks, whereas the addition of 3-hydroxybutyrate decreased 14CO2 production from lactate only in synaptic terminals from 17- to 18-day-old rat brain. None of the added substrates altered the rate of 14CO2 production from labelled glutamine or malate suggesting that these substrates are metabolized in relatively distinct compartments within synaptic terminals. Overall the data demonstrate that synaptic terminals from both weanling and adult rat brain can utilize a variety of substrates for energy. In addition, the competition studies demonstrate that the interactions of substrates change with age and suggest that there are multiple compartments of energy metabolism (or tricarboxylic acid cycle activity) in isolated synaptic terminals.

Asunto(s)

RESUMEN

We evaluated ictal 99mtechnetium hexamethyl propylene-amine-oxime single-photon emission computed tomography (SPECT) in 22 children with electroclinical features of frontal lobe epilepsy (FLE). Ictal SPECT demonstrated unilateral frontal hyperperfusion in 20 of 22 children (91%) (one lobar, two frontocentral, six dorsolateral, six frontopolar, three orbitofrontal, one medial frontal, and one insula), concordant with electroclinical lateralization in 19 of 20 (95%). Hyperperfusion was evident in the ipsilateral basal ganglia in 16 of 22 (73%) and the contralateral cerebellum in 14 of 22 children (64%). Interictal SPECT showed unilateral, localized frontal hypoperfusion concordant with electroclinical lateralization in only two of 22 children (9%). Ictal SPECT localization to the frontocentral, media frontal, or dorsolateral regions was associated with asymmetric tonic posturing, contralateral head/eye deviation, and unilateral clonic jerking (p < 0.01). Ictal SPECT localization to the frontopolar or orbitofrontal regions was associated with vocalization, hyperventilation, truncal flexion, and complex gestural automatisms (p > or = 0.05). Ictal SPECT has the potential to (1) localize seizures in patients with intractable FLE, and (2) advance understanding of the in vivo anatomico-clinical relationships of frontal lobe seizures.

Asunto(s)

RESUMEN

Seventeen ictal 99mTc-HMPAO single photon emission computed tomography (SPECT) studies were performed in 15 children with temporal lobe epilepsy (TLE) aged 7-14 years (mean 10.3 years). Ictal SPECT was informative in 16 of 17 (94%) studies in 14 of 15 (93%) children, showing unilateral temporal lobe hyperperfusion. In all 16 informative ictal SPECT studies, lateralization was concordant with ictal EEG, magnetic resonance imaging (MRI), and pathology. In 4 children, ictal SPECT provided additional localizing information that was not apparent from concurrent ictal EEG recording. Blinded interpretation of ictal SPECT studies by two independent investigators showed correct lateralization of the epileptic focus in every child. Results of visual analysis of ictal SPECT images were corroborated by quantitative analysis. Although interictal SPECT studies showed a degree of temporal lobe hypoperfusion in all children, in 9 of 15 hypoperfusion was either minimal, bilateral, contralateral, or associated with extratemporal hypoperfusion. In children with TLE, ictal SPECT provides reliable lateralizing information to corroborate or supplement that obtained from surface EEG and MRI.

Asunto(s)

RESUMEN

We reviewed the outcome of corpus callosal section in 64 adult and pediatric patients to identify factors associated with a good outcome: 48% of patients had a favorable outcome for overall seizure frequency. Improvement was noted in several seizure types and was most likely for drop attacks, particularly in the setting of a unilateral focal cerebral lesion or a true generalized epilepsy of Lennox-Gastaut type. Poor outcomes for drop attacks were more likely if there was associated severe intellectual handicap or bilateral independent spikes on interictal EEG. Complex partial seizures (CPS), most commonly of frontal lobe origin, also responded favorably. The complications of callosal section were usually mild and transient. New focal seizures occurred in only 2 patients and were not as frequent or disabling as preoperative seizures types. A worthwhile improvement in seizure outcome was achieved by completion of the callosotomy in 6 of 10 patients with unsatisfactory results from anterior callosotomy.

Asunto(s)

RESUMEN

This is the first published report of Landau-Kleffner syndrome occurring discordantly in monozygotic twins. The implications of this finding in the understanding of the etiology of this condition are discussed. Five other patients are reported and comparisons are drawn with other series.

Asunto(s)