Hypothalamic pregnenolone mediates recognition memory in the context of metabolic disorders

Obesity and type 2 diabetes are associated with cognitive dysfunction. Because the hypothalamus is implicated in energy balance control and memory disorders, we hypothesized that specific neurons in this brain region are at the interface of metabolism and cognition. Acute obesogenic diet administration in mice impaired recognition memory due to defective production of the neurosteroid precursor pregnenolone in the hypothalamus. Genetic interference with pregnenolone synthesis by Star deletion in hypothalamic POMC, but not AgRP neurons, deteriorated recognition memory independently of metabolic disturbances. Our data suggest that pregnenolone’s effects on cognitive function were mediated via an autocrine mechanism on POMC neurons, influencing hippocampal long-term potentiation. The relevance of central pregnenolone on cognition was also confirmed in metabolically unhealthy patients with obesity. Our data reveal an unsuspected role for POMC neuron-derived neurosteroids in cognition. These results provide the basis for a framework to investigate new facets of POMC neuron biology with implications for cognitive disorders.

Elucidating the role of neuron-astrocyte metabolic coupling in learning and memory

Monika Tadi

The brain has very high energy demands that are mainly met by the circulating blood glucose to ensure its proper functioning. Thus, it is not surprising that though the human brain weighs only 2- 3% of the body weight, it consumes approximately 25% of total body glucose. “Neuro-energetic coupling" is a unique feature of the brain and refers to the existence of the tight coupling between (neuronal) synaptic brain activity and energy metabolism. The main excitatory neurotransmitter in the brain is glutamate and most of the energy requirements at the cortical level are met through glutamate-mediated neurotransmission, suggesting that there is a close relationship between brain activity, glutamatergic neurotransmission and energy metabolism. Until the 80s, blood-borne glucose was thought to be the sole energy substrate of the brain cells and it was assumed that glucose is metabolized by neurons and astrocytes (a type of glial cell) independently. It is only during the last two decades that the concept of compartmentalization of energy metabolism between neurons and astrocytes has become more obvious. The transfer of lactate from astrocytes to neurons termed as the “astrocyte-neuron lactate shuttle (ANLS)” model proposed by Dr. Pellerin and Prof. Magistretti posits that glutamate released during brain activation activates glycolysis (one of glucose metabolic pathways) in astrocytes leading to lactate (a monocarboxylate) release in the extracellular space, and this astrocytic lactate is then captured and used as an energy substrate by the activated neurons to meet their energy requirements. It includes a complex chain of events involved in neuro-metabolic coupling and supports the preferential use of lactate by activated neurons under conditions of increased energy demands. Higher brain functions such as learning a new task are associated with structural changes in brain and are metabolically demanding necessitating the need of an additional energy supply to meet the neurons increased energy requirement. In this context, the current thesis project aimed to elucidate the contribution of metabolic coupling between astrocytes and neurons, known as “neuron-astrocyte metabolic coupling” to learning and memory formation. Using an in vivo approach combined with behavioral, molecular and gene manipulation techniques in mice, we set out to investigate the role of neuron-astrocyte metabolic coupling, particularly ANLS in cognition. The project was divided into two main parts: In the first part, the expression of glucose metabolism-related genes was investigated during long term memory (LTM) formation in fear-motivated inhibitory avoidance (IA) learning paradigm. Using (14C) 2-Deoxyglucose (2-DG) technique, we first defined the brain areas that are involved in IA LTM formation. This technique provides an indirect measurement of brain activity by measuring glucose uptake in different brain areas during the task. The results of brain metabolic mapping show an increase in glucose utilization in the hippocampus, amygdala, anterior cingulate cortex and pre-limbic and infra-limbic cortical areas. Results from the quantitative analysis of mRNA levels in the dorsal hippocampus at different time point’s following IA task highlight a time dependent, learning specific change in the expression of genes related to glucose metabolism. Specific set of genes involved in the synthesis and degradation of glycogen (brain glucose reserve present only in astrocytes), pyruvate metabolism and pentose phosphate shunt as well as the ANLS were induced following the IA task. These early observations indicate that the metabolic interactions between astrocytes and neurons undergo modifications during a learning process and suggest that these adaptations may play a key role in the establishment of long-term memory. In the second part of research project, we focused on investigating the behavioral consequences of down regulating the expression of Monocarboxylate Transporter 1 (MCT1), a key ANLS related gene, on cognition and higher brain functions. Mice heterozygous for the MCT1 gene (MCT1+/-) were characterized in numerous behavioral paradigms such as general wellbeing, reflexive and motor capabilities. After having established that MCT1+/- mice have no alterations in general emotional state or in locomotion, a battery of behavioral tests was performed to study cognition in these mice. Compared with wild-type control mice, the MCT1+/- mice display normal muscle strength and motor coordination. However, when tested for higher brain functions, the MCT1+/- mice exhibit learning and memory deficits in several learning paradigms/tasks that are hippocampus and / or amygdala dependent. The cognitive impairment observed in the MCT1 heterozygous mice further highlights the importance of ANLS in memory and suggest that disruption of lactate transfer from astrocytes to neurons via the MCT1 results in cognitive impairment. The overall results in this thesis demonstrate that cerebral energy metabolism, and in particular the transfer of lactate from astrocytes to neurons not only provides temporal adaptations in the learning process, but it also plays a fundamental role in memory formation.

EPFL2013

The Impact of Sleep Fragmentation on Sleep Homeostasis, Brain and Peripheral Energy Metabolism and Spatial Learning

Maxime Olivier Baud

The quality of sleep has recently come to the forefront of public health concerns in industrialized nations. Indeed, voluntary sleep curtailment is widespread, sleep disorders are increasingly recognized and both correlate with the current epidemiology of diabetes and obesity (Van Cauter and Knutson, 2008). Sleep fragmentation (SF) is a periodic disruption found in highly prevalent sleep-related breathing and movement disorders, where the sleeper might be aroused several hundred times per night. By roughly summing the prevalence of obstructive sleep apnea (OSA) syndromes and periodic limb movement disorders, two of the most common sleep disorders, we predict a total prevalence for SF of nearly 15% in the European active population. Overlooked cases and insufficient concern for the problematic bear a major socioeconomic burden because of the broad impact of excessive daytime sleepiness on productivity (Hossain and Shapiro, 2002), motor vehicle accidents (Aldrich, 1989) and medical complications (Spiegel et al., 2009). Recurrent interference with the natural architecture of sleep leads to sleepiness, difficulty in concentration and cognitive impairments such as decreased reaction times and poor working memory equivalent to a full night of sleep deprivation in severe cases (Bonnet, 1985, 1989; Stepanski, 2002). In addition, recent experimental evidence in humans has unraveled the unprecedented link between sleep of poor quality and dramatic endocrine imbalances such as deregulation of appetite controlling hormones, glucocorticoids and increase in sympathetic tone, ultimately leading to the development of insulin resistance (Spiegel et al., 2009; Stamatakis and Punjabi, 2010). Current animal models for the study of sleep pathology mostly involve short intervention often biased by stress induction and usually involving a complete period of sleep deprivation. These models do not tightly reflect the most common clinical pattern characterized by sleep perturbation over long periods of time. Therefore, paying attention to maximally avoid methodological stress, we designed a new device aiming at performing instrumental SF for fourteen days in mice. Our model mimics SF observed in sleep disorders since it has no major impact on circadian timing and total amount of sleep but clearly shortens individual sleep episodes. With this method, we approached three problematics that were insufficiently addressed in the literature. In our first study, we addressed the question of whether SF could trigger electrophysiological and molecular sleep homeostasis mechanisms in the short-term and whether these effects could be maintained in the long-term. We showed that SF leads to an anticipated homeostatic regulation of Slow Wave Activity (SWA), the sleep EEG hallmark of sleep homeostasis. Furthermore, EEG spectral analysis revealed an unexpected power increase in 4-40 Hz frequencies during sleep with a clear increase in the amplitude of slow waves and spindles. We concluded that sustained sleep pressure in SF leads to a global increase in cortical synchronization. We next explored whether these electrophysiological changes could impact on the expression of transcripts known to be regulated by sleep homeostasis (immediate early genes, clock-genes and heat shock proteins). Only heat shock proteins showed a consistent induction after one day but normalized after fourteen days of SF. These results point to the fact that electrophysiological and molecular aspects of sleep homeostasis are dissociable and that long-term sleep pressure triggers allostatic cellular processes of adaptation. In our second study, we explored the effects of long-term SF on general metabolism. We showed that SF led to a general imbalance in energy intake and expenditure, possibly through a slight elevation of brain temperature at rest. We also observed that glucocorticoids were slightly elevated at given circadian points and were accompanied by glucose intolerance. Altogether, this study brings evidence for the participation of SF in addition to intermittent hypoxia in the development of a metabolic syndrome in OSA patients. In the third study, we combined our chronic model with a spatial learning task spanning over several days (Morris Water Maze) in order to study the impact of SF on memory and metabolic plasticity. We showed that SF impaired proper memory consolidation supporting the idea that continuous sleep is required for specific mechanisms of memory trace post-processing. We also provide preliminary evidence that brain energy metabolism is not optimally responsive to neural activation after SF and that this effect might underlie the observed cognitive impairment. In conclusion, we showed that our animal SF model is suitable for studying the specific mechanisms underlying cognitive impairments and metabolic imbalance observed in sleep medicine. Our results underscore the existence of a global allostatic load in a wide range of systems during SF. Sensitization of the population to the prevalence of SF and its stealthy mode of presentation might improve its detection. Increased recognition of the problematic would be especially beneficial to cognitive development in school children with sleep apnea and might reduce accidents related to sleepiness. More specifically, we propose that screening sleep quality and effective correction of SF in patients with endocrine disorders has the potential to improve glucose metabolism and energy balance.

EPFL2011

Impact of individual differences in glucocorticoid adaptation to stress on behavior, neurophysiology and metabolism

Damien Sébastien Huzard

The stress system is a key modulator of homeostasis and allows organisms to adapt to environmental changes. Proper survival is dependent on the appropriate stress response, for example initiating food (energy) intake or provoking physical reaction. However, long-term activity of the stress system is related to cardiovascular diseases, metabolic syndromes as well as accelerated aging and cognitive impairments. In order to assess the impacts of stress regulation on cardiac, metabolic and aging health processes, we used lines of rats selected for their differential glucocorticoid responsiveness to stress during juvenile period, in the different experiments presented here. The three lines of rats showed low, intermediate or high response to stress and elicited differences in biobehavioral phenotypes. Cardiovascular diseases are highly exacerbated by stress exposure and autonomic imbalance. Reduced vagal modulation has been related to a lower stress flexibility and deleterious effects on cardiac health. In a first study, we investigated the autonomic nervous system modulation of heart rate in the three lines of rats (differing in their responsiveness to stress). Electrocardiographic recordings were performed at rest and following autonomic pharmacological manipulations. Rats with intermediate reactivity to stress had a higher resting parasympathetic (vagal) modulation and a reduced heart rate compared to rats with low or high stress responses. Furthermore, pharmacological treatments showed that the sympathetic regulation of the heart was not impaired in rats with low and high responsiveness to stress. Stress can affect social interactions and, in return, social interactions can be the cause of critical stress. Furthermore, since the stress system is related to key metabolic mediators, we investigated in a second study, the general metabolism of rats from the three lines. Moreover, we paired rats from the different lines together, in mixed-line dyads, and we evaluated the differences in social interactions and the long-term effects of mixed-line pairing on metabolism. We used indirect calorimetry and mitochondrial respirometry to measure energy expenditure and mitochondrial function. We observed that the selection for differences in glucocorticoid responsiveness induced constitutive differences in energy expenditure and fuel use. Moreover, we showed that the biobehavioral phenotypes affected the social interactions between the different lines. Finally, long-term mixed-line pairing affected global and central metabolism of the rats, with rats from the low and intermediate responsive lines being more susceptible to changes. In a final experiment, we studied the interaction of two risk factors for cognitive decline, the secretion of corticosterone and aging. Indeed, dysfunctions of the stress system contribute and facilitate aging and increased glucocorticoids induce cognitive alterations. We assessed anxiety, stress responsiveness, coping-style and cognitive functions in a Morris water-maze at early-aging. Results indicated that the phenotype of the lines were stable throughout life and that learning, swimming strategies and reversal ability were different between the lines. Overall, we showed that this model is suitable to study the systems related to stress regulation. Future research may use this animal model in order to investigate further the relationship between opposite stress regulation and health.

EPFL2018