In vitro measurement of enzymatic markers as a tool to detect mouse cardiomyocytes injury

Primary cultures of cardiomyocytes represent a useful model for analyzing cardiac cell biology as well as pathogenesis of several cardiovascular disorders. Our aim was to standardize protocols for determining the damage of cardiac cells cultured in vitro by measuring the creatine kinase and its cardiac isotype and lactate dehydrogenase activities in the supernatants of mice cardiomyocytes submitted to different protocols of cell lysis. Our data showed that due to its higher specificity, the cardiac isotype creatine kinase was the most sensitive as compared to the others studied enzymatic markers, and can be used to monitor and evaluate cardiac damage in in vitro assays.

Brain aging in the oldest-old.

Nonagenarians and centenarians represent a quickly growing age group worldwide. In parallel, the prevalence of dementia increases substantially, but how to define dementia in this oldest-old age segment remains unclear. Although the idea that the risk of Alzheimer's disease (AD) decreases after age 90 has now been questioned, the oldest-old still represent a population relatively resistant to degenerative brain processes. Brain aging is characterised by the formation of neurofibrillary tangles (NFTs) and senile plaques (SPs) as well as neuronal and synaptic loss in both cognitively intact individuals and patients with AD. In nondemented cases NFTs are usually restricted to the hippocampal formation, whereas the progressive involvement of the association areas in the temporal neocortex parallels the development of overt clinical signs of dementia. In contrast, there is little correlation between the quantitative distribution of SP and AD severity. The pattern of lesion distribution and neuronal loss changes in extreme aging relative to the younger-old. In contrast to younger cases where dementia is mainly related to severe NFT formation within adjacent components of the medial and inferior aspects of the temporal cortex, oldest-old individuals display a preferential involvement of the anterior part of the CA1 field of the hippocampus whereas the inferior temporal and frontal association areas are relatively spared. This pattern suggests that both the extent of NFT development in the hippocampus as well as a displacement of subregional NFT distribution within the Cornu ammonis (CA) fields may be key determinants of dementia in the very old. Cortical association areas are relatively preserved. The progression of NFT formation across increasing cognitive impairment was significantly slower in nonagenarians and centenarians compared to younger cases in the CA1 field and entorhinal cortex. The total amount of amyloid and the neuronal loss in these regions were also significantly lower than those reported in younger AD cases. Overall, there is evidence that pathological substrates of cognitive deterioration in the oldest-old are different from those observed in the younger-old. Microvascular parameters such as mean capillary diameters may be key factors to consider for the prediction of cognitive decline in the oldest-old. Neuropathological particularities of the oldest-old may be related to "longevity-enabling" genes although little or nothing is known in this promising field of future research.

Neurochemical profile of the developing mouse cortex determined by in vivo 1H NMR spectroscopy at 14.1 T and the effect of recurrent anaesthesia.

The neurochemical profile of the cortex develops in a region and time specific manner, which can be distorted by psychiatric and other neurological pathologies. Pre-clinical studies often involve experimental mouse models. In this study, we determined the neurochemical profile of C57BL/6 mice in a longitudinal study design to provide a reference frame for the normal developing mouse cortex. Using in vivo proton NMR spectroscopy at 14 T, we measured the concentrations of 18 metabolites in the anterior and posterior cortex on postnatal days (P) 10, 20, 30, 60 and 90. Cortical development was marked by alterations of highly concentrated metabolites, such as N-acetylaspartate, glutamate, taurine and creatine. Regional specificity was represented by early variations in the concentration of glutamine, aspartate and choline. In adult animals, regional concentration differences were found for N-acetylaspartate, creatine and myo-inositol. In this study, animals were exposed to recurrent isoflurane anaesthesia. Additional experiments showed that the latter was devoid of major effects on behaviour or cortical neurochemical profile. In conclusion, the high sensitivity and reproducibility of the measurements achieved at 14 T allowed us to identify developmental variations of cortical areas within the mouse cortex.

Sleep loss reduces the DNA-binding of BMAL1, CLOCK, and NPAS2 to specific clock genes in the mouse cerebral cortex.

We have previously demonstrated that clock genes contribute to the homeostatic aspect of sleep regulation. Indeed, mutations in some clock genes modify the markers of sleep homeostasis and an increase in homeostatic sleep drive alters clock gene expression in the forebrain. Here, we investigate a possible mechanism by which sleep deprivation (SD) could alter clock gene expression by quantifying DNA-binding of the core-clock transcription factors CLOCK, NPAS2, and BMAL1 to the cis-regulatory sequences of target clock genes in mice. Using chromatin immunoprecipitation (ChIP), we first showed that, as reported for the liver, DNA-binding of CLOCK and BMAL1 to target clock genes changes in function of time-of-day in the cerebral cortex. Tissue extracts were collected at ZT0 (light onset), -6, -12, and -18, and DNA enrichment of E-box or E'-box containing sequences was measured by qPCR. CLOCK and BMAL1 binding to Cry1, Dbp, Per1, and Per2 depended on time-of-day, with maximum values reached at around ZT6. We then observed that SD, performed between ZT0 and -6, significantly decreased DNA-binding of CLOCK and BMAL1 to Dbp, consistent with the observed decrease in Dbp mRNA levels after SD. The DNA-binding of NPAS2 and BMAL1 to Per2 was also decreased by SD, although SD is known to increase Per2 expression in the cortex. DNA-binding to Per1 and Cry1 was not affected by SD. Our results show that the sleep-wake history can affect the clock molecular machinery directly at the level of chromatin binding thereby altering the cortical expression of Dbp and Per2 and likely other targets. Although the precise dynamics of the relationship between DNA-binding and mRNA expression, especially for Per2, remains elusive, the results also suggest that part of the reported circadian changes in DNA-binding of core clock components in tissues peripheral to the suprachiasmatic nuclei could, in fact, be sleep-wake driven.

Protein kinase C-activating tumor promoters enhance the differentiation of astrocytes in aggregating fetal brain cell cultures.

Serum-free aggregating cell cultures of fetal rat telencephalon treated with the potent tumor promoter phorbol 12-myristate 13-acetate (PMA) showed a marked, rapid, and sustained increase in the activity of the astrocyte-specific enzyme glutamine synthetase (GS). This effect was accompanied by a small increase in RNA synthesis and a progressive reduction in DNA synthesis. Only mitotically active cultures were responsive to PMA treatments. Since in aggregate cultures astrocytes are the preponderant cell type, both in number and mitotic activity, it can be concluded that PMA induces and/or enhances the terminal differentiation of astrocytes. The developmental expression of GS was also greatly stimulated by mezerein, a potent nonphorbol tumor promoter, but not by 4 alpha-phorbol 12,13-didecanoate, a nonpromoting phorbol ester. Since both tumor promoters, PMA and mezerein, are potent and specific activators of C-kinase, it is suggested that C-kinase plays a regulatory role in the growth and differentiation of normal astrocytes.

The pattern and Loci of training-induced brain changes in healthy older adults are predicted by the nature of the intervention.

There is enormous interest in designing training methods for reducing cognitive decline in healthy older adults. Because it is impaired with aging, multitasking has often been targeted and has been shown to be malleable with appropriate training. Investigating the effects of cognitive training on functional brain activation might provide critical indication regarding the mechanisms that underlie those positive effects, as well as provide models for selecting appropriate training methods. The few studies that have looked at brain correlates of cognitive training indicate a variable pattern and location of brain changes - a result that might relate to differences in training formats. The goal of this study was to measure the neural substrates as a function of whether divided attentional training programs induced the use of alternative processes or whether it relied on repeated practice. Forty-eight older adults were randomly allocated to one of three training programs. In the SINGLE REPEATED training, participants practiced an alphanumeric equation and a visual detection task, each under focused attention. In the DIVIDED FIXED training, participants practiced combining verification and detection by divided attention, with equal attention allocated to both tasks. In the DIVIDED VARIABLE training, participants completed the task by divided attention, but were taught to vary the attentional priority allocated to each task. Brain activation was measured with fMRI pre- and post-training while completing each task individually and the two tasks combined. The three training programs resulted in markedly different brain changes. Practice on individual tasks in the SINGLE REPEATED training resulted in reduced brain activation whereas DIVIDED VARIABLE training resulted in a larger recruitment of the right superior and middle frontal gyrus, a region that has been involved in multitasking. The type of training is a critical factor in determining the pattern of brain activation.

Cerebral extracellular lactate increase is predominantly nonischemic in patients with severe traumatic brain injury.

Growing evidence suggests that endogenous lactate is an important substrate for neurons. This study aimed to examine cerebral lactate metabolism and its relationship with brain perfusion in patients with severe traumatic brain injury (TBI). A prospective cohort of 24 patients with severe TBI monitored with cerebral microdialysis (CMD) and brain tissue oxygen tension (PbtO2) was studied. Brain lactate metabolism was assessed by quantification of elevated CMD lactate samples (>4 mmol/L); these were matched to CMD pyruvate and PbtO2 values and dichotomized as glycolytic (CMD pyruvate >119 μmol/L vs. low pyruvate) and hypoxic (PbtO2 <20 mm Hg vs. nonhypoxic). Using perfusion computed tomography (CT), brain perfusion was categorized as oligemic, normal, or hyperemic, and was compared with CMD and PbtO2 data. Samples with elevated CMD lactate were frequently observed (41±8%), and we found that brain lactate elevations were predominantly associated with glycolysis and normal PbtO2 (73±8%) rather than brain hypoxia (14±6%). Furthermore, glycolytic lactate was always associated with normal or hyperemic brain perfusion, whereas all episodes with hypoxic lactate were associated with diffuse oligemia. Our findings suggest predominant nonischemic cerebral extracellular lactate release after TBI and support the concept that lactate may be used as an energy substrate by the injured human brain.

Dual role of Bmi1 loss in the preservation of photoreceptor layers in the Rd1 mouse

Purpose: In the Rd1 and Rd10 mouse models of retinitis pigmentosa, a mutation in the Pde6ß gene leads to the rapid loss of photoreceptors. As in several neurodegenerative diseases, Rd1 and Rd10 photoreceptors re-express cell cycle proteins prior to death. Bmi1 regulates cell cycle progression through inhibition of CDK inhibitors, and its deletion efficiently rescues the Rd1 retinal degeneration. The present study evaluates the effects of Bmi1 loss in photoreceptors and Müller glia, since in lower vertebrates, these cells respond to retinal injury through dedifferentiation and regeneration of retinal cells. Methods: Cell death and Müller cell activation were analyzed by immunostaining of wild-type, Rd1 and Rd1;Bmi1-/- eye sections during retinal degeneration, between P10 and P20. Lineage tracing experiments use the GFAP-Cre mouse (JAX) to target Müller cells. Results: In Rd1 retinal explants, inhibition of CDKs reduces the amount of dying cells. In vivo, Bmi1 deletion reduces CDK4 expression and cell death in the P15 Rd1;Bmi1-/- retina, although cGMP accumulation and TUNEL staining are detected at the onset of retinal degeneration (P12). This suggests that another process acts in parallel to overcome the initial loss of Rd1;Bmi1-/- photoreceptors. We demonstrate here that Bmi1 loss in the Rd1 retina enhances the activation of Müller glia by downregulation of p27Kip1, that these cells migrate toward the ONL, and that some cells express the retinal progenitor marker Pax6 at the inner part of the ONL. These events are also observed, but to a lesser extent, in Rd1 and Rd10 retinas. At P12, EdU incorporation shows proliferating cells with atypical elongated nuclei at the inner border of the Rd1;Bmi1-/- ONL. Lineage tracing targeting Müller cells is in process and will determine the implication of this cell population in the maintenance of the Rd1;Bmi1-/- ONL thickness and whether downregulation of Bmi1 in Rd10 Müller cells equally stimulates their activation. Conclusions: Our results show a dual role of Bmi1 deletion in the rescue of photoreceptors in the Rd1;Bmi1-/- retina. Indeed, the loss of Bmi1 reduces Rd1 retinal degeneration, and as well, enhances the Müller glia activation. In addition, the emergence of cells expressing a retinal progenitor marker in the ONL suggests Bmi1 as a blockade to the regeneration of retinal cells in mammals.

Vasopressin-stimulated CFTR Cl- currents are increased in the renal collecting duct cells of a mouse model of Liddle's syndrome.

Liddle's syndrome is a genetic form of hypertension linked to Na(+) retention caused by activating mutations in the COOH terminus of the beta or gamma subunit of the epithelial sodium channel (ENaC). In this study, we used the short-circuit current (I(sc)) method to investigate the effects of deamino-8-d-arginine vasopressin (dDAVP) on Na(+) and Cl(-) fluxes in primary cultures of cortical collecting ducts (CCDs) microdissected from the kidneys of mice with Liddle's syndrome carrying a stop codon mutation, corresponding to the beta-ENaC R(566) stop mutation (L) found in the original pedigree. Compared to wild-type (+/+) CCD cells, untreated L/+ and L/L CCD cells exhibited 2.7- and 4.2-fold increases, respectively, in amiloride-sensitive (Ams) I(sc), reflecting ENaC-dependent Na(+) absorption. Short-term incubation with dDAVP caused a rapid and significant increase (approximately 2-fold) in Ams I(sc) in +/+, but not in L/+ or L/L CCD cells. In sharp contrast, dDAVP induced a greater increase in 5-nitro-2-(3-phenylpropamino)benzoate (NPPB)-inhibited apical Cl(-) currents in amiloride-treated L/L and L/+ cells than in their +/+ counterparts. I(sc) recordings performed under apical ion substituted conditions revealed that the dDAVP-stimulated apical secretion of Cl(-), which was absent in cultured CCDs lacking CFTR, was 1.8-fold greater in L/+ and 3.7-fold greater in L/L CCD cells than in their +/+ CCD counterparts. After the basal membrane had been permeabilized with nystatin and a basal-to-apical Cl(-) gradient had been imposed, dDAVP also stimulated larger Cl(-) currents across L/L and L/+ CCD layers than +/+ CCD layers. These findings demonstrate that vasopressin stimulates greater apical CFTR Cl(-) conductance in the renal CCD cells of mice with Liddle's syndrome than in wild-type mice. This effect could contribute to the enhanced NaCl reabsorption observed in the distal nephron of patients with Liddle's syndrome.

MOLECULAR REORGANIZATION FOLLOWING SENSORY STIMULATION IN THE ADULT MOUSE BARREL CORTEX

Sensory information is an important factor in shaping neuronal circuits during development and adulthood. In the barrel cortex of adult rodents, cells from layer IV are able to adapt their functional state to an increased flow of sensory information from the mystacial whisker follicles. Previous studies in our group have shown that whisker stimulation induces the formation of inhibitory synapses in the corresponding barrel (Knott et al., 2002) and decreases neuronal responses toward the deflection of the stimulated whisker (Quairiaux et al., 2007). Together these observations have turned the barrel cortex into a model to study homeostatic plasticity. At the cellular level, neuronal activity triggers intracellular signaling cascades leading to a transcriptional response. To further characterize the molecular pathways involved in the synaptic changes after whisker stimulation in the adult mouse, a previous doctoral student in our group performed a microarray analysis on laser-dissected barrels in sections through layer IV. This study identified the regulation (up and down) of a series of genes in the stimulated barrels (thesis of Johnston-Wenger, 2010). We here focused on ten genes that presented the highest fold change according to the microarray analysis. Out of these genes, 7 are known as neuronal activity-dependent genes (Tnncl, Nptx2, Sorcs3, Ptgs2, Nr4a2, Npas4 and Adcyapl) whereas three have so far not been related to neuronal plasticity (Scn7a, Pcdhl5 and Cede3). The study aimed at confirming the results of the microarray analysis and localizing molecular modifications in the stimulated barrel column at the cellular level. In situ hybridization for Pcdhl5 after different periods of whisker stimulation (3, 6, 9, 15, 24 hrs) allowed us to confirm that the 1.25 fold change used for the microarray analysis is an appropriate threshold for considering a regulation significant after sensory-stimulation. Moreover, we confirmed with in situ hybridization a significant upregulation of the genes of interest in the stimulated barrels. In situ hybridization and immunohistochemistry allowed us to observe the distribution of the genes of interest and the corresponding protein products at the cellular level. Three observations were made: 1) alterations of the expression was restricted to the stimulated barrels for all genes tested; 2) within a barrel column not all cells responded to whisker stimulation with an altered gene expression; 3) in the stimulated barrels, two different patterns of mRNA and protein expression can be distinguished. We hypothesize that this segregation of the activity-induced gene expression reflects the segregation of the two principal thalamocortical pathways conveying the sensory information to the barrel cortex. Moreover, only neurons reaching the critical threshold will modify their gene expression program resulting in structural as well as physiological modifications that prevent the subsequent propagation of the excess of excitation to the postsynaptic targets. The activity-induced gene expression is therefore adapted in a cell-type-specific manner to induce a homeostatic response to the entire neuronal network involved in the integration of the sensory information. This to our knowledge the first study showing the distinct, but complementary contribution of the two thalamocortical pathways in experience-dependent plasticity in the adult mouse barrel cortex. -- L'information sensorielle nous permet de continuellement façonner nos circuits neuronaux autant durant le développement qu'à l'âge adulte. Chez le rongeur l'information sensorielle perçue par les vibrisses est intégrée au niveau du cortex somatosensoriel primaire (appelé en anglais « barrel cortex ») dont les cellules de la couche IV sont capables d'adapter leur état fonctionnel en réponse à une augmentation d'activité neuronale. Ce modèle expérimental a permis à notre groupe de recherche d'observer des changements rapides du circuit neuronal en fonction de l'activité sensorielle. En effet, la stimulation continue d'une vibrisse d'une souris adulte pendant 24 heures induit non seulement un remaniement synaptique (Knott et al., 2002), mais également des changements physiologiques au niveau des neurones du tonneau correspondant (Quairiaux et al., 2007). Ces observations nous permettent d'affirmer que le « barrel cortex » est un modèle approprié pour y étudier la plasticité synaptique. Au niveau cellulaire, l'activité neuronale déclenche des cascades de signalisation intracellulaire résultant en une réponse transcriptionnelle. Afin de caractériser les voies moléculaires impliquées dans la plasticité synaptique, une puce à ARN nous a permis de comparer l'expression de gènes entre un tonneau correspondant à une vibrisse stimulée et un tonneau d'une vibrisse non-stimulée (Nathalie). Cette analyse a révélé un certain nombre de gènes régulés de manière positive ou négative par l'augmentation de l'activité neuronale. Nous nous sommes concentrés sur 10 gènes dont l'expression est fortement régulée. L'expression de sept d'entre eux a déjà été démontrée comme dépendante de l'activité neuronale (Tnncl, Nptx2, Sorcs3, Ptgs2, Nr4a2, Npas4 otAdcyapl) alors que l'expression des trois autres (Scn7a, Pcdhl5 et Cedei) n'a pour le moment pas encore été liée à la plasticité neuronale. Le but de cette thèse est de confirmer les résultats de la puce à ARN et de déterminer dans quel type cellulaire ces gènes sont exprimés. L'hybridation in situ pour le gène Pcdhl5, après différentes périodes de stimulation des vibrisses (3, 6, 9, 15 et 24 heures), nous a permis de confirmer que le seuil de 1.25x utilisé dans l'analyse de la puce à ARN est approprié pour considérer qu'un gène est régulé de manière significative par la stimulation sensorielle. Nous avons également pu confirmer à l'aide de cette technique que la stimulation sensorielle augmente significativement l'expression de ces dix gènes. L'expression de ces gènes au niveau cellulaire a été observée à l'aide des techniques d'hybridation in situ et d'immunohistochimie. Trois observations ont été faites : 1) la régulation de ces gènes est restreinte aux tonneaux correspondants aux vibrisses stimulées ; 2) au niveau d'une colonne corticale correspondant aux vibrisses stimulées, seules certaines cellules présentent une altération de leur expression génique ; 3) au niveau des tonneaux stimulés, deux profils d'expression d'ARNm et de protéines sont observés. Notre hypothèse est que cette distribution pourrait correspondre à la terminaison ségrégée des deux voies thalamocortical qui amènent l'information sensorielle dans le cortex cérébral. De plus, seul les neurones atteignant le seuil critique d'activation modifient leur expression génique en réponse à la stimulation sensorielle. Ces changements d'expression géniques vont permettre à la cellule de modifier ses propriétés structurales et physiologiques de manière a prevenir la propagation d'un excès d'activité neuronale au niveau de ses cibles postsynaptics. L'activité neuronale agit donc spécifiquement sur certains types cellulaires de maniere a induire une réponse homéostatique au niveau du réseau neuronal impliqué dans l'integration de l'information sensorielle. Nos travaux démontrent pour une première fois que les deux voies sensorielles contribuent d'une manière distincte et complémentaire à la plasticité corticale induite par un changement de l'activité sensorielle chez la souris adulte.

Influence of epidermal growth factor on the maturation of fetal rat brain cells in aggregate culture. An immunocytochemical study.

Maturation of astrocytes, neurons, and oligodendrocytes was studied in serum-free aggregating cell cultures of fetal rat telencephalon by an immunocytochemical approach. Cell type-specific immunofluorescence staining was examined by using antibodies directed against glial fibrillary acidic protein (GFAP) and vimentin, two astroglial markers; neuron-specific enolase (NSE) and neurofilament (NF), two neuronal markers, and galactocerebroside (GC), an oligodendroglial marker. It was found that the cellular maturation in aggregates is characterized by distinct developmental increases in immunoreactivity for GFAP, vimentin, NSE, NF, and GC, and by a subsequent decrease of vimentin-positive structures in more differentiated cultures. These findings are in agreement with observations in vivo, and they corroborate previous biochemical studies of this histotypic culture system. Treatment of very immature cultures with a low dose of epidermal growth factor (EGF, 5 ng/ml) enhanced the developmental increase in GFAP, NSE, NF and GC immunoreactivity, suggesting an acceleration of neuronal and glial maturation. In addition, EGF was found to alter the cellular organization within the aggregates, presumably by influencing cell migration.

Immunocytochemical localization of the glucose transporter 2 (GLUT2) in the adult rat brain. II. Electron microscopic study.

Following a former immunohistochemical study in the rat brain [Arluison, M., Quignon, M., Nguyen, P., Thorens, B., Leloup, C., Penicaud, L. Distribution and anatomical localization of the glucose transporter 2 (GLUT2) in the adult rat brain. I. Immunohistochemical study. J. Chem. Neuroanat., in press], we have analyzed the ultrastructural localization of GLUT2 in representative and/or critical areas of the forebrain and hindbrain. In agreement with previous results, we observe few oligodendrocyte and astrocyte cell bodies discretely labeled for GLUT2 in large myelinated fibre bundles and most brain areas examined, whereas the reactive glial processes are more numerous and often localized in the vicinity of nerve terminals and/or dendrites or dendritic spines forming synaptic contacts. Only some of them appear closely bound to unlabeled nerve cell bodies and dendrites. Furthermore, the nerve cell bodies prominently immunostained for GLUT2 are scarce in the brain nuclei examined, whereas the labeled dendrites and dendritic spines are relatively numerous and frequently engaged in synaptic junctions. In conformity with the observation of GLUT2-immunoreactive rings at the periphery of numerous nerve cell bodies in various brain areas (see previous paper), we report here that some neuronal perikarya of the dorsal endopiriform nucleus/perirhinal cortex exhibit some patches of immunostaining just below the plasma membrane. However, the presence of many GLUT2-immunoreactive nerve terminals and/or astrocyte processes, some of them being occasionally attached to nerve cell bodies and dendrites, could also explain the pericellular labeling observed. The results here reported support the idea that GLUT2 may be expressed by some cerebral neurones possibly involved in glucose sensing, as previously discussed. However, it is also possible that this transporter participate in the regulation of neurotransmitter release and, perhaps, in the release of glucose by glial cells.

Brain dysfunction in sepsis: what can we learn from cerebral perfusion studies?

Investigations on the relationship between sepsis, brain dysfunction, and cerebral perfusion are methodologically very difficult to perform. It is important to interpret the results of such studies in view of our limited ability to diagnose and quantify brain dysfunction and to consider our limited understanding of the mechanisms that lead to or are associated with brain dysfunction in sepsis.

Low plasma lactate concentration as a biomarker of an incompetent brain-pull: a risk factor for weight gain in type 2 diabetes patients.

OBJECTIVE: Body weight development is closely regulated by central nervous mechanisms. As has been demonstrated recently, the capability of the brain to actively demand energy from the body (brain-pull) is indispensable for the maintenance of systemic homeostasis. A deficit in this brain-pull may result in compensatory ingestive behavior followed by weight gain in the medium or long term. The aim of this study was to establish a biomarker of such an incompetent brain-pull. Since lactate is an alternative cerebral energy substrate to glucose, we investigated whether low fasting plasma lactate concentrations are associated with weight gain and increased feelings of hunger in patients with type 2 diabetes over a 3-year period. METHODS: In a population based cohort study 134 type 2 diabetes patients were examined at baseline and 3-year follow-up. Plasma lactate concentrations and additional hormones associated with food intake such as e.g. insulin, or leptin, as well as psychological variables like hunger feelings before and after a standardized breakfast were measured. The relation between fasting plasma lactate concentrations and postprandial hunger as well as follow-up weight was analyzed. RESULTS: Low fasting plasma lactate concentrations predicted a higher 3-year follow-up weight (B=-1.268, SE=0.625, p=0.04). Moreover, low fasting plasma lactate concentrations were associated with more pronounced feelings of postprandial hunger (B=-0.406, SE=0.137, p<0.01). CONCLUSIONS: We conclude that low plasma lactate concentrations may represent a biomarker of an incompetent brain-pull, which is associated with weight gain and increased postprandial hunger in patients with type 2 diabetes mellitus. These results are in line with the view that plasma lactate can be used by the brain as an alternative energy substrate and thereby to some extent prevent overeating and obesity.