Activation of Stat3 Signaling in AgRP Neurons Promotes Locomotor Activity

Over the last years, much of the research on obesity has focused on the study of leptin. This adipocyte-derived hormone circulates in proportion to fat mass and functions as an adiposity signal to decrease energy intake and increase energy expenditure in order to maintain energy homeostasis. Leptin signals informations on body energy stores to hypothalamic neurons located in the arcuate nucleus (ARC) of the hypothalamus. One of the leptin-regulated neuronal subtypes in the ARC are the orexigenic agouti-related peptide (AgRP)-producing neurons, which are directly inhibited by leptin. A key pathway downstream of the leptin receptor involves activation of the signal transducer and activator of transcription 3 (Stat3), but the role of Stat3 in the regulation of AgRP neurons remains controversial. In this study, analysis of Stat3-CAgRP mice expressing a constitutively active version of the Stat3 protein (Stat3-C) selectively in AgRP neurons reveals a crucial role for Stat3 in AgRP neurons in the regulation of energy expenditure in vivo. Stat3-CAgRP mice are lean and develop a relative resistance to diet-induced obesity accompanied by improved glucose homeostasis. The lean phenotype of Stat3-CAgRP mice appears in the presence of unaltered AgRP expression and caloric intake as a consequence of increased energy expenditure evoked by elevated locomotor activity. Consistent with the phenotype observed in Stat3-CAgRP mice, expression of Stat3-C in AgRP neurons of leptin deficient ob/ob mice diminishes the obese phenotype of ob/ob mice as a result of increased energy expenditure and locomotor activity in the presence of unaltered food intake. Analysis of brain catecholamines in Stat3-CAgRP mice revealed a trend towards elevated dopamine concentrations in the striatum and frontal cortex, which potentially account for the increased locomotor activity in those mice. Nevertheless, the anatomical interaction of AgRP neurons with neuronal centers that control locomotor activity and the exact molecular mechanism in AgRP neurons leading to Stat3-dependent activation of locomotor activity have to be defined further. Taken together, this thesis introduces a novel model according to which leptin-stimulated Stat3 activation in AgRP neurons directly regulates locomotor activity independent of the regulation of AgRP mRNA expression

Preserved respiratory chain capacity and physiology in mice with profoundly reduced levels of mitochondrial respirasomes

The mammalian respiratory chain complexes I, III 2, and IV (CI, CIII 2, and CIV) are critical for cellular bioenergetics and form a stable assembly, the respirasome (CI-CIII 2-CIV), that is biochemically and structurally well documented. The role of the respirasome in bioenergetics and the regulation of metabolism is subject to intense debate and is difficult to study because the individual respiratory chain complexes coexist together with high levels of respirasomes. To critically investigate the in vivo role of the respirasome, we generated homozygous knockin mice that have normal levels of respiratory chain complexes but profoundly decreased levels of respirasomes. Surprisingly, the mutant mice are healthy, with preserved respiratory chain capacity and normal exercise performance. Our findings show that high levels of respirasomes are dispensable for maintaining bioenergetics and physiology in mice but raise questions about their alternate functions, such as those relating to the regulation of protein stability and prevention of age-associated protein aggregation

Sex-specific effects of Cre expression in Syn1Cre mice

Abstract The Cre-loxP system has been used to generate cell-type specific mutations in mice, allowing researchers to investigate the underlying biological mechanisms of disease. However, the Cre-recombinase alone can induce phenotypes that confound comparisons among genotypes if the appropriate Cre control is not included. In this study, we characterised behavioural, morphological and metabolic phenotypes of the pan-neuronal Syn1Cre line. We found that these mice possess intact neuromuscular parameters but have reduced exploratory activity and a male-specific increase in anxiety-like behaviour. Moreover, we observed a male-specific deficit in learning and long-term memory of Syn1Cre mice that could be a result of decreased visual acuity. Furthermore, we found that over-expression of human growth hormone (hGH) from Syn1Cre results in a male-specific reduction in body weight and femur length, potentially through decreased hepatic Igf1 expression. However, metabolic characteristics of Syn1Cre mice such as glucose metabolism, energy expenditure and feeding were unaffected by the presence of Syn1Cre. In conclusion, our data demonstrate that Syn1Cre expression has effects on behavioural and morphological traits. This finding highlights the importance of including the Cre control in all comparisons, while the male-specific effects on some phenotypes highlight the importance of including both sexes

Hexosamine pathway activation improves memory but does not extend lifespan in mice

Glucosamine feeding and genetic activation of the hexosamine biosynthetic pathway (HBP) have been linked to improved protein quality control and lifespan extension. However, as an energy sensor, the HBP has been implicated in tumor progression and diabetes. Given these opposing outcomes, it is imperative to explore the long-term effects of chronic HBP activation in mammals. Thus, we asked if HBP activation affects metabolism, coordination, memory, and survival in mice. N-acetyl-D-glucosamine (GlcNAc) supplementation in the drinking water had no adverse effect on weight in males but increased weight in young females. Glucose or insulin tolerance was not affected up to 20 months of age. Of note, we observed improved memory in young male mice supplemented with GlcNAc. Survival was not changed by GlcNAc treatment. To assess the effects of genetic HBP activation, we overexpressed the pathway's key enzyme GFAT1 and a constitutively activated mutant form in all mouse tissues. We detected elevated levels of the HBP product UDP-GlcNAc in mouse brains, but did not find any effects on behavior, memory, or survival. Together, while dietary GlcNAc supplementation did not extend survival in mice, it positively affected memory and is generally well tolerated

High levels of TFAM repress mammalian mitochondrial DNA transcription in vivo

Mitochondrial transcription factor A (TFAM) is compacting mitochondrial DNA (dmtDNA) into nucleoids and directly controls mtDNA copy number. Here, we show that the TFAM-to-mtDNA ratio is critical for maintaining normal mtDNA expression in different mouse tissues. Moderately increased TFAM protein levels increase mtDNA copy number but a normal TFAM-to-mtDNA ratio is maintained resulting in unaltered mtDNA expression and normal whole animal metabolism. Mice ubiquitously expressing very high TFAM levels develop pathology leading to deficient oxidative phosphorylation (OXPHOS) and early postnatal lethality. The TFAM-to-mtDNA ratio varies widely between tissues in these mice and is very high in skeletal muscle leading to strong repression of mtDNA expression and OXPHOS deficiency. In the heart, increased mtDNA copy number results in a near normal TFAM-to-mtDNA ratio and maintained OXPHOS capacity. In liver, induction of LONP1 protease and mitochondrial RNA polymerase expression counteracts the silencing effect of high TFAM levels. TFAM thus acts as a general repressor of mtDNA expression and this effect can be counterbalanced by tissue-specific expression of regulatory factors

Dnmt3a2/Dnmt3L Overexpression in the Dopaminergic System of Mice Increases Exercise Behavior through Signaling Changes in the Hypothalamus

Dnmt3a2, ade novoDNA methyltransferase, is induced by neuronal activity and participates in long-term memory formation with the increased expression of synaptic plasticity genes. We wanted to determine if Dnmt3a2 with its partner Dnmt3L may influence motor behavior via the dopaminergic system. To this end, we generated a mouse line, Dnmt3a2/3L(Dat/wt), with dopamine transporter (DAT) promotor driven Dnmt3a2/3L overexpression. The mice were studied with behavioral paradigms (e.g., cylinder test, open field, and treadmill), brain slice patch clamp recordings, ex vivo metabolite analysis, and in vivo positron emission tomography (PET) using the dopaminergic tracer 6-[F-18]FMT. The results showed that spontaneous activity and exercise performance were enhanced in Dnmt3a2/3L(Dat/wt)mice compared to Dnmt3a2/3L(wt/wt)controls. Dopaminergic substantia nigra pars compacta neurons of Dnmt3a2/3L(Dat/wt)animals displayed a higher fire frequency and excitability. However, dopamine concentration was not increased in the striatum, and dopamine metabolite concentration was even significantly decreased. Striatal 6-[F-18]FMT uptake, reflecting aromatic L-amino acid decarboxylase activity, was the same in Dnmt3a2/3L(Dat/wt)mice and controls. [F-18]FDG PET showed that hypothalamic metabolic activity was tightly linked to motor behavior in Dnmt3a2/3L(Dat/wt)mice. Furthermore, dopamine biosynthesis and motor-related metabolic activity were correlated in the hypothalamus. Our findings suggest that Dnmt3a2/3L, when overexpressed in dopaminergic neurons, modulates motor performance via activation of the nigrostriatal pathway. This does not involve increased dopamine synthesis

Hippocampal insulin resistance links maternal obesity with impaired neuronal plasticity in adult offspring

Objective: Maternal obesity and a disturbed metabolic environment during pregnancy and lactation have been shown to result in many long-term health consequences for the offspring. Among them, impairments in neurocognitive development and performance belong to the most dreaded ones. So far, very few mechanistic approaches have aimed to determine the responsible molecular events. Methods: In a mouse model of maternal diet-induced obesity and perinatal hyperinsulinemia, we assessed adult offspring's hippocampal insulin signaling as well as concurrent effects on markers of hippocampal neurogenesis, synaptic plasticity and function using western blotting and immunohistochemistry. In search for a potential link between neuronal insulin resistance and hippocampal plasticity, we additionally quantified protein expression of key molecules of synaptic plasticity in an in vitro model of acute neuronal insulin resistance. Results: Maternal obesity and perinatal hyperinsulinemia result in adult hippocampal insulin resistance with subsequently reduced hippocampal mTor signaling and altered expression of markers of neurogenesis (doublecortin), synaptic plasticity (FoxO1, pSynapsin) and function (vGlut, vGAT) in the offspring. The observed effects are independent of the offspring's adult metabolic phenotype and can be associated with multiple previously reported behavioral abnormalities. Additionally, we demonstrate that induction of insulin resistance in cultured hippocampal neurons reduces mTor signaling, doublecortin and vGAT protein expression. Conclusions: Hippocampal insulin resistance might play a key role in mediating the long-term effects of maternal obesity and perinatal hyperinsulinemia on hippocampal plasticity and the offspring's neurocognitive outcome

Preserved respiratory chain capacity and physiology in mice with profoundly reduced levels of mitochondrial respirasomes.

The mammalian respiratory chain complexes I, III2, and IV (CI, CIII2, and CIV) are critical for cellular bioenergetics and form a stable assembly, the respirasome (CI-CIII2-CIV), that is biochemically and structurally well documented. The role of the respirasome in bioenergetics and the regulation of metabolism is subject to intense debate and is difficult to study because the individual respiratory chain complexes coexist together with high levels of respirasomes. To critically investigate the in vivo role of the respirasome, we generated homozygous knockin mice that have normal levels of respiratory chain complexes but profoundly decreased levels of respirasomes. Surprisingly, the mutant mice are healthy, with preserved respiratory chain capacity and normal exercise performance. Our findings show that high levels of respirasomes are dispensable for maintaining bioenergetics and physiology in mice but raise questions about their alternate functions, such as those relating to the regulation of protein stability and prevention of age-associated protein aggregation

Matching Dietary Amino Acid Balance to the In Silico-Translated Exome Optimizes Growth and Reproduction without Cost to Lifespan

Balancing the quantity and quality of dietary protein relative to other nutrients is a key determinant of evolutionary fitness. A theoretical framework for defining a balanced diet would both reduce the enormous workload to optimize diets empirically and represent a breakthrough toward tailoring diets to the needs of consumers. Here, we report a simple and powerful in silico technique that uses the genome information of an organism to define its dietary amino acid requirements. We show for the fruit fly Drosophila melanogaster that such "exome-matched" diets are more satiating, enhance growth, and increase reproduction relative to non-matched diets. Thus, early life fitness traits can be enhanced at low levels of dietary amino acids that do not impose a cost to lifespan. Exome matching also enhanced mouse growth, indicating that it can be applied to other organisms whose genome sequence is known