LEC 14 Hypothalamic Control of Food Intake PDF

LEC 14: Hypothalamic Control of Food Intake Since 1975, global obesity rates have tripled, affecting over one-third of the world's population by 2016, with approximately 650 million individuals classified as clinically obese. The obesity epidemic is not limited to adults; there is a significant incr...

LEC 14: Hypothalamic Control of Food Intake Since 1975, global obesity rates have tripled, affecting over one-third of the world's population by 2016, with approximately 650 million individuals classified as clinically obese. The obesity epidemic is not limited to adults; there is a significant increase in childhood obesity, with 41 million children under the age of five being overweight in 2016. Obesity is commonly assessed using the Body Mass Index (BMI), calculated by dividing weight in kilograms by height in square meters. A BMI between 25 and 30 indicates overweightness, while a BMI exceeding 30 signifies clinical obesity; despite LEC 14: Hypothalamic Control of Food Intake 1 BMI's imperfections, it remains widely utilized. The depiction of obesity rates in the United States on a map illustrates a concerning trend, with each state color-coded to represent the percentage of individuals with a BMI over 30. This transition from predominantly yellow (BMI 10 to 20) in 1990 to increasing shades of red and blue indicates a substantial rise in obesity prevalence, with projections suggesting further escalation by 2030 if current trends persist. The expanding obesity epidemic is not unique to the United States but is a characteristic trend in developed countries globally, indicating a complex and widespread public health issue. The rise in obesity is influenced by various factors including socio-economic disparities, cultural norms, environmental influences, and genetic predispositions. The expansion of the obesity epidemic can be attributed to our obesogenic environment, characterized by increased food consumption and larger portion sizes over time. Despite being recognized as unhealthy, soda remains popular due to its palatability, leading to excessive consumption, as evidenced by data from the Coca-Cola website comparing calorie content with fruit juice. While both soda and fruit juice may contain similar calorie amounts per volume, the palatability of soda encourages greater consumption, a phenomenon not observed with less palatable options like fruit juice. Sedentary lifestyles also play a crucial role in obesity, as modern routines involve less physical activity, exacerbating weight gain over time. Obesity is associated with various chronic diseases, including type 2 diabetes, cardiovascular diseases, hypertension, hyperlipidemia, sleep disorders, fertility issues, and certain types of cancer, underscoring its detrimental impact on health and well-being. LEC 14: Hypothalamic Control of Food Intake 2 Body weight regulation adheres to the first law of thermodynamics, where energy intake must equal energy expenditure to maintain equilibrium; any disparity leads to weight changes. The brain governs appetite through integration of various peripheral and neural signals, with two primary systems controlling body weight: the homeostatic and hedonic systems. 1. The homeostatic system, akin to a thermostat, adjusts energy balance by modulating hunger and eating behavior based on energy stores; decreased energy stores trigger hunger signals, prompting food intake. Brain structures involved in these regulatory mechanisms include the hypothalamus and brainstem for the homeostatic system, with the hypothalamus serving as a central hub for homeostatic control. The hypothalamus, located at the base of the brain, serves as a primary regulator of appetite and energy balance. Mouse models are commonly utilized in studying appetite regulation, with brain sections providing insights into hypothalamic structure and function. Sagittal sections divide the brain into right and left, while coronal sections separate front and back regions, aiding in the visualization of hypothalamic nuclei. Early lesioning experiments in the 1940s identified the hypothalamus as a key site for energy homeostasis regulation, with targeted destruction of specific hypothalamic nuclei leading to alterations in body weight. 1. Lesions in the ventromedial nuclei, dorsomedial nuclei, or paraventricular nuclei resulted in obesity in animal models, highlighting the importance of these regions in appetite regulation. Arc is also important shown below 2. Conversely, lesions in the lateral hypothalamic area (LHA) led to decreased food intake and subsequent weight loss or anorexia in animals. LEC 14: Hypothalamic Control of Food Intake 3 2. In contrast, the hedonic system operates independently of hunger and energy status, responding to pleasure sensations associated with food consumption, potentially overriding the homeostatic pathway. Hedonic or reward-based eating can lead to overconsumption, particularly in environments abundant with palatable food, contributing to rising obesity rates, with the hedonic system playing a significant role in this phenomenon. An illustrative example of hedonic system dominance occurs when individuals, despite feeling full after a meal, indulge in dessert purely for pleasure, disregarding energy intake considerations, showcasing how hedonic impulses override homeostatic signals. The ventral tegmental area (VTA), in the midbrain, serves as the main hub for the hedonic system. Two popular mouse models, the ob/ob and db/db mice, emerged as valuable tools for understanding body weight regulation due to their spontaneous mutations resulting in obesity. Compared to the typical adult wild-type mouse weighing between 25 and 35 grams, ob/ob and db/db mice can reach weights of up to 100 grams, illustrating their obesity phenotype. Dr. Douglas Coleman conducted parabiosis experiments in the 1970s, surgically joining ob/ob or db/db mice with wild-type mice to explore the role of circulating factors in appetite control. When ob/ob mice were paired with wild-type mice, they exhibited reduced food intake and eventually reached a normal body weight, LEC 14: Hypothalamic Control of Food Intake 4 suggesting the presence of an appetite-suppressing factor in wild-type circulation and a receptor for that factor in the ob/ob. In contrast, db/db mice paired with wild-type mice did not experience a similar reduction in body weight, indicating that db/db mice lacked the receptor for the appetite-suppressing factor provided by the wild type. When ob/ob and db/db mice were paired together, ob/ob exhibited decreased food intake and eventually died from starvation, implying that the ob/ob had the receptor for the appetite-suppressing factor produced by and acquired from the db/db mice. In 1994, Dr. Jeffrey Friedman's laboratory at Rockefeller University successfully sequenced the gene responsible for the obesity phenotype observed in ob/ob mice, naming it leptin from the Greek word "leptos," meaning thin. Leptin, the hormone identified as deficient in ob/ob mice, is primarily produced in adipocytes or fat cells, with its secretion regulated by the size and number of adipocytes. Leptin is released into circulation and acts as a signal to the brain, communicating information about energy stores and regulating appetite and metabolism accordingly. Subsequent research, two years later, confirmed that db/db mice were deficient in the leptin receptor, further elucidating the role of leptin signaling in appetite regulation and obesity. Dr. Jeffrey Friedman and Dr. Douglas Coleman, pictured here, played pivotal roles in these groundbreaking discoveries, expanding our understanding of the genetics underlying obesity. Their contributions were recognized with prestigious awards, including the Albert Lasker Award in 2010, an honor often associated with Nobel Prize winners, underscoring the significance of their work in the field of obesity research. LEC 14: Hypothalamic Control of Food Intake 5 Treating ob/ob mice, deficient in leptin, with leptin effectively reduces their body weight, demonstrating the therapeutic potential of leptin in combating obesity. Similar positive outcomes were observed in humans diagnosed with leptin deficiency after the gene sequencing breakthrough in 1994, with individuals experiencing significant weight normalization upon daily leptin treatment. Initially hailed as a promising anti-obesity drug, leptin's efficacy was questioned as subsequent studies revealed that most obese individuals are not leptin deficient; instead, they exhibit hyperleptinemia, characterized by elevated circulating leptin levels and leptin resistance. Leptin resistance, wherein the body fails to respond to leptin signals despite high levels of circulating leptin, remains poorly understood and is believed to develop gradually over time. As individuals age and gain weight, leptin sensitivity diminishes, contributing to the development of leptin resistance, although the precise mechanisms underlying this phenomenon remain elusive. Despite ongoing research efforts, including investigations in the professor's laboratory, elucidating the cellular and molecular mechanisms of leptin resistance and its implications remains a complex challenge in the field of neuroendocrinology. Leptin binds to its receptor, causing the receptor to dimerize, initiating the leptin signaling cascade. An important component of this signaling pathway is the phosphorylation of STAT3 (signal transducer and activator of transcription 3) >> a transcription factor, which serves as an indicator of leptin activity. Phosphorylated STAT3 is frequently used as a marker to assess the effectiveness of leptin signaling. The arcuate nucleus of the hypothalamus is a critical brain area for energy balance and leptin sensitivity. Leptin levels increase with food intake, signaling the body's energy status to the brain, particularly to neurons in the arcuate nucleus. There are two main types of neurons in the arcuate nucleus: POMC (pro-opiomelanocortin) neurons, which decrease appetite. LEC 14: Hypothalamic Control of Food Intake 6 NPY/AgRP (neuropeptide Y/agouti-related protein) neurons, which increase appetite. Leptin activates POMC neurons and inhibits NPY/AgRP neurons, leading to a reduction in appetite. Ghrelin, a hormone released from the gastrointestinal tract, opposes leptin's inhibitory action by activating NPY/AgRP neurons, thus increasing appetite. The interaction between leptin and ghrelin on these neurons provides a simplified model of how the brain regulates food intake through signaling pathways. Neurosurgery has been a foundational tool in studying the neurobiology of appetite control, particularly through lesioning experiments in the mid-20th century to understand brain functions. Immunohistochemistry is another crucial technique, using antibodies to detect brain proteins under various conditions, such as fasting or obesity, to study their impact on energy homeostasis. The discovery that levels of AgRP increase in the fasted state compared to the fed state highlighted its role in appetite control. This was evidenced by immunohistochemical staining, where a fed mouse's brain showed less AgRP (red signal) compared to a fasted mouse's brain, which displayed a significant increase in the red signal. Similar findings were observed with NPY, which also showed increased levels in a fasted state, suggesting its role in promoting feeding and increased appetite. To determine if NPY and AgRP co-localize within the same neurons, double immunohistochemistry was employed. The co-localization would result in a yellow or orange signal if both proteins were present in the same neurons, confirming that NPY and AgRP are expressed by the same neuronal population. Another neuropeptide, POMC (pro-opiomelanocortin), was LEC 14: Hypothalamic Control of Food Intake 7 identified as anorexigenic, reducing feeding when administered to mice. Investigating its co-expression with NPY and AgRP showed no overlap, indicating distinct neuronal populations within the arcuate nucleus. Genetic manipulation, especially Cre-loxP recombination, serves as a crucial method in neurobiology for exploring the roles of hormones and proteins by facilitating site-specific DNA recombination. Site-specific Cre recombinase enzyme activates DNA recombination at the loxP sites which possess unique sequences enabling precise genetic alterations. This approach is instrumental for dissecting gene functions in designated tissues or cells by positioning the Cre recombinase gene under the control of a specific promoter. In the Cre-loxP system, the Cre recombinase enzyme recognizes and interacts with loxP sites, which are specific 34base pair DNA sequences. These loxP sites are artificially inserted flanking the gene of interest (on both the 5' and 3' sides) within the genome of an organism or in a plasmid. When the Cre enzyme is expressed in the cell, it recombines the DNA at the loxP sites, effectively excising the gene located between them. This leads to the disruption of the gene's function, either by removing it entirely from the genome or by making it non-functional. When the Cre recombinase acts on two loxP sites that are oriented in the same direction on a DNA molecule, the DNA sequence flanked by these loxP sites is excised or removed. This is often used to delete a gene or a specific DNA sequence to study its function by observing the effects of its absence. For recombination, the orientation of the loxP sites influences the outcome of the recombination event: If the loxP sites are oriented in opposite directions, the Cre recombinase will invert the DNA sequence flanked by these sites rather than excising it. This can be used to control gene expression or to replace a gene with another. To examine a protein's role in the liver, for instance, researchers position the Cre recombinase gene (which codes for the Cre enzyme) under a liver-specific promoter, ensuring targeted gene disruption solely in liver cells. Similarly, if you want to express the Cre enzyme in the brain, you would employ a neuron specific promoter. A significant application of this method was in studying leptin signaling, employing Cre-loxP recombination to ascertain the leptin receptor's significance in distinct tissues. The db/db mouse model, characterized by a universal absence of leptin receptors, led scientists to investigate leptin's action sites—whether it operates locally in adipocytes or elsewhere. Targeting neuronal tissue, scientists employed neuron-specific promoters for Cre recombinase expression, selectively abolishing the leptin receptor in LEC 14: Hypothalamic Control of Food Intake 8 the brain. Observations from mice with brain-specific leptin receptor deletions, which exhibited severe obesity akin to db/db mice, affirmed that leptin signaling predominantly transpires in the brain. Researchers use GFP (green fluorescent protein) under the leptin receptor promoter to visualize leptin receptor-expressing neurons in the mouse brain, with each expressing neuron turning green. The highest density of leptin receptor-expressing neurons, indicated by the brightest green signal, is located in the arcuate nucleus of the hypothalamus, suggesting its critical role in leptin signaling. To identify key neuronal populations in leptin signaling, researchers systematically knocked out the leptin receptor in specific neuron populations. Knocking out the leptin receptor in POMC neurons resulted in mice that were heavier than controls, but the difference in body weight was relatively modest, approximately 5 grams. Similar experiments on neurons expressing AgRP did not yield dramatic changes in body weight, indicating no significant impact on obesity when leptin receptors were removed from these neurons alone. Further studies involved knocking out the leptin receptor in neurons of the ventromedial hypothalamus, which also did not lead to significant weight differences compared to the control. A combined knockout of leptin receptors in both POMC and ventromedial hypothalamic neurons showed an additive effect on weight gain, yet still did not replicate the substantial obesity observed in db/db mice. The conclusion is that while LEC 14: Hypothalamic Control of Food Intake 9 knocking out leptin receptors in specific neuronal populations can lead to mild obesity, only the complete absence of leptin receptors throughout the brain, as seen in db/db mice, results in massive obesity. This indicates a distributed role of leptin signaling in regulating body weight. Researchers divided neurons into GABAergic (inhibitory) and glutamatergic (excitatory) populations and observed the effects of leptin receptor knockout in each group. Mice lacking leptin receptors in GABAergic neurons exhibited obesity comparable to db/db mice, indicating a critical role of GABAergic neurons in mediating leptin's effects on body weight. In contrast, mice with leptin receptor knockout in glutamatergic neurons were overweight but not to the extent of those lacking receptors in GABAergic neurons, highlighting the differential impact of these neuronal types on leptin signaling and body weight regulation. This finding underscores the importance of GABAergic neurons in leptin's anorectic (appetite-reducing) actions, suggesting that leptin primarily influences appetite and weight through inhibitory neuronal pathways. LEC 14: Hypothalamic Control of Food Intake 10 To link neuronal activity with feeding behavior, researchers utilized a calcium sensor, a mutated GFP fused with calmodulin, to visualize neuronal activity in real-time. The sensor brightens in response to calcium binding, indicating neuronal activity. Focusing on AgRP neurons, known for their role in promoting feeding, the study employed a miniature microscope to observe these neurons' activity in live mice during feeding behavior experiments. Observations showed that AgRP neuronal activity decreased when food was introduced, suggesting that these neurons play a role in food-seeking behavior rather than directly measuring the body's energy status. The decrease in neuronal activity occurred even when non-edible objects resembling food were introduced, indicating that the neurons respond to the potential of food availability rather than its actual consumption. This study reveals that AgRP neurons are primarily involved in the motivation for food seeking, with their activity diminishing upon the discovery of food or food-like objects, suggesting a complex interplay between neuronal activity, feeding behavior, and energy homeostasis. Optogenetics, employing channel rhodopsin, a light-sensitive ion channel, allows for the manipulation of neuron activity. When neurons equipped with channel rhodopsin are illuminated with blue light, they facilitate the transport of cations, notably calcium, into the cell, thereby modulating cellular activity. In this context, the technique was specifically directed at AgRP-expressing neurons within the arcuate nucleus. By placing channel rhodopsin under the control of the AgRP promoter, researchers ensured that only these neurons, critical for hunger signaling, were targeted for activation. LEC 14: Hypothalamic Control of Food Intake 11 Activating AgRP neurons optogenetically by shining blue light resulted in an immediate increase in appetite behaviors in mice, with the animals moving towards food sources. This direct intervention illustrates the causal relationship between AgRP neuron activity and the initiation of feeding behavior. Further elucidating the influence of AgRP neurons on behavior, a study employed optogenetic activation during a flavor preference experiment. Mice were presented with two non-caloric food options, one flavored with strawberry and the other with orange, with neither flavor inherently preferred by the mice. During the conditioning phase, blue light activation of AgRP neurons was coupled with the consumption of the orange-flavored food, embedding a negative association with this flavor through the induced hunger sensation. Preference testing later revealed that mice developed a distinct aversion to the orange flavor, avoiding it in favor of the strawberry flavor, which had not been associated with the optogenetically induced hunger sensation. This outcome demonstrated that AgRP neuron activation not only triggers hunger but also can generate a lasting aversion to specific stimuli associated with its activation. These results highlight the profound impact of AgRP neurons on not just the physiological aspect of hunger but also on the complex behaviors related to food preference and aversion, underscoring the aversive nature of hunger and the psychological challenges posed by dieting. LEC 14: Hypothalamic Control of Food Intake 12