Interoception and the origin of feelings: A new synthesis.

Feelings are conscious mental events that represent body states as they undergo homeostatic regulation. Feelings depend on the interoceptive nervous system (INS), a collection of peripheral and central pathways, nuclei and cortical regions which continuously sense chemical and anatomical changes in the organism. How such humoral and neural signals come to generate conscious mental states has been a major scientific question. The answer proposed here invokes (1) several distinctive and poorly known physiological features of the INS; and (2) a unique interaction between the body (the 'object' of interoception) and the central nervous system (which generates the 'subject' of interoception). The atypical traits of the INS and the direct interactions between neural and non-neural physiological compartments of the organism, neither of which is present in exteroceptive systems, plausibly explain the qualitative and subjective aspects of feelings, thus accounting for their conscious nature.

The 4E-BP growth pathway regulates the effect of ambient temperature on Drosophila metabolism and lifespan.

Changes in body temperature can profoundly affect survival. The dramatic longevity-enhancing effect of cold has long been known in organisms ranging from invertebrates to mammals, yet the underlying mechanisms have only recently begun to be uncovered. In the nematode Caenorhabditis elegans, this process is regulated by a thermosensitive membrane TRP channel and the DAF-16/FOXO transcription factor, but in more complex organisms the underpinnings of cold-induced longevity remain largely mysterious. We report that, in Drosophila melanogaster, variation in ambient temperature triggers metabolic changes in protein translation, mitochondrial protein synthesis, and posttranslational regulation of the translation repressor, 4E-BP (eukaryotic translation initiation factor 4E-binding protein). We show that 4E-BP determines Drosophila lifespan in the context of temperature changes, revealing a genetic mechanism for cold-induced longevity in this model organism. Our results suggest that the 4E-BP pathway, chiefly thought of as a nutrient sensor, may represent a master metabolic switch responding to diverse environmental factors.

The nature of feelings: evolutionary and neurobiological origins.

Feelings are mental experiences of body states. They signify physiological need (for example, hunger), tissue injury (for example, pain), optimal function (for example, well-being), threats to the organism (for example, fear or anger) or specific social interactions (for example, compassion, gratitude or love). Feelings constitute a crucial component of the mechanisms of life regulation, from simple to complex. Their neural substrates can be found at all levels of the nervous system, from individual neurons to subcortical nuclei and cortical regions.

Quantifying Drosophila food intake: comparative analysis of current methodology.

Food intake is a fundamental parameter in animal studies. Despite the prevalent use of Drosophila in laboratory research, precise measurements of food intake remain challenging in this model organism. Here, we compare several common Drosophila feeding assays: the capillary feeder (CAFE), food labeling with a radioactive tracer or colorimetric dye and observations of proboscis extension (PE). We show that the CAFE and radioisotope labeling provide the most consistent results, have the highest sensitivity and can resolve differences in feeding that dye labeling and PE fail to distinguish. We conclude that performing the radiolabeling and CAFE assays in parallel is currently the best approach for quantifying Drosophila food intake. Understanding the strengths and limitations of methods for measuring food intake will greatly advance Drosophila studies of nutrition, behavior and disease.

Water- and nutrient-dependent effects of dietary restriction on Drosophila lifespan.

Dietary restriction (DR) is a widely conserved intervention leading to lifespan extension. Despite considerable effort, the mechanisms underlying DR remain poorly understood. In particular, it remains unclear whether DR prolongs life through conserved mechanisms in different species. Here, we show that, in the most common experimental conditions, lifespan extension by DR is abolished by providing Drosophila with ad libitum water, without altering food intake, indicating that DR, as conventionally studied in flies, is fundamentally different from the phenomenon studied in mammals. We characterize an alternative dietary paradigm that elicits robust lifespan extension irrespective of water availability, and thus likely represents a more relevant model for mammalian DR. Our results support the view that protein:carbohydrate ratio is the main dietary determinant of fly lifespan. These findings have broad implications for the study of lifespan and nutrition.

Allocrine modulation of feeding behavior by the Sex Peptide of Drosophila.

Mating elicits a dramatic reprogramming of female behavior in numerous insect species. In Drosophila, this postmating response (PMR) comprises increased egg-laying rate and reduced sexual receptivity and is controlled by the products of the male accessory glands, a family of approximately 80 small peptides transferred in the male seminal fluid . Here, we show that copulation strongly stimulates female food intake. Remarkably, this change is abolished if the males lack a single, small seminal protein, the Sex Peptide (SP). Ectopic expression of SP in virgin females mimics the effect of mating on feeding behavior, demonstrating that SP is the main agent controlling this behavioral paradigm. Our observations identify enhanced feeding behavior as a novel component of the Drosophila PMR and suggest that SP represents a molecular link between energy acquisition and reproductive investment.

A Role for The P2Y1 Receptor in Nonsynaptic Cross-depolarization in the Rat Dorsal Root Ganglia.

Non-synaptic transmission is pervasive throughout the nervous system. It appears especially prevalent in peripheral ganglia, where non-synaptic interactions between neighboring cell bodies have been described in both physiological and pathological conditions, a phenomenon referred to as cross-depolarization (CD) and thought to play a role in sensory processing and chronic pain. CD has been proposed to be mediated by a chemical agent, but its identity has remained elusive. Here, we report that in the rat dorsal root ganglion (DRG), the P2Y1 purinergic receptor (P2RY1) plays an important role in regulating CD. The effect of P2RY1 is cell-type specific: pharmacological blockade of P2RY1 inhibited CD in A-type neurons while enhancing it in C-type neurons. In the nodose ganglion of the vagus, CD requires extracellular calcium in a large percentage of cells. In contrast, we show that in the DRG extracellular calcium appears to play no major role, pointing to a mechanistic difference between the two peripheral ganglia. Furthermore, we show that DRG glial cells also play a cell-type specific role in CD regulation. Fluorocitrate-induced glial inactivation had no effect on A-cells but enhanced CD in C-cells. These findings shed light on the mechanism of CD in the DRG and pave the way for further analysis of non-synaptic neuronal communication in sensory ganglia.

From membrane excitability to metazoan psychology.

Unlike the nonexcitable cell membranes that are ubiquitous in all domains of life, excitable membranes are found almost exclusively in animal organisms (Protozoa and Metazoa). Their transient permeability to ion flow makes possible the rapid detection of, and response to, external stimuli, and results in the phenomena that most clearly distinguish fauna from flora: perception, cognition, and motor activity. Interestingly, all known forms of membrane excitability are a consequence of one unique mechanism: the influx of positively charged ions into the normally alkaline cytoplasm. Here, we suggest that the sudden reversal of the membrane potential during the sensory potential and the action potential is an electrostatic disturbance of homeostasis that is the necessary first step in the processes of 'sentience' and 'irritability'.

High carbohydrate-low protein consumption maximizes Drosophila lifespan.

Dietary restriction extends lifespan in a variety of organisms, but the key nutritional components driving this process and how they interact remain uncertain. In Drosophila, while a substantial body of research suggests that protein is the major dietary component affecting longevity, recent studies claim that carbohydrates also play a central role. To clarify how nutritional factors influence longevity, nutrient consumption and lifespan were measured on a series of diets with varying yeast and sugar content. We show that optimal lifespan requires both high carbohydrate and low protein consumption, but neither nutrient by itself entirely predicts lifespan. Increased dietary carbohydrate or protein concentration does not always result in reduced feeding-the regulation of food consumption is best described by a constant daily caloric intake target. Moreover, due to differences in food intake, increased concentration of a nutrient within the diet does not necessarily result in increased consumption of that particular nutrient. Our results shed light on the issue of dietary effects on lifespan and highlight the need for accurate measures of nutrient intake in dietary manipulation studies.

Non-lethal PCR genotyping of single Drosophila.

In Drosophila, genetic techniques relying on stochastic chromosomal rearrangements involve the generation and screening of a large number of fly stocks to isolate a few lines of interest. Here, we describe a PCR-based method allowing non-lethal molecular characterization of single flies. Using this procedure, individual candidate recombinant animals can be genotyped and selected one generation earlier than with extant methodology and, importantly, before stocks are established. This advance should significantly facilitate several of the most fundamental and routine techniques in Drosophila genetics.

The Drosophila G protein-coupled receptor, Methuselah, exhibits a promiscuous response to peptides.

Methuselah (Mth) is a G protein-coupled receptor (GPCR) associated with longevity in Drosophila melanogaster. Previously, Stunted (Sun) was identified as a peptide agonist of Mth. Here, we identify two additional activators of Mth signaling: Drosophila Sex Peptide (SP) and a novel peptide (Serendipitous Peptide Activator of Mth, SPAM). Minimal functional sequences and key residues were identified from Sun and SPAM by studying truncation and alanine-scanning mutations. These peptide agonists share little sequence homology and illustrate the promiscuity of Mth for activation. mth mutants exhibit no defects in behaviors controlled by SP, casting doubt on the biological significance of Mth activation by any of these agonists, and illustrating the difficulty in applying in vitro studies to their relevance in vivo. Future studies of Mth ligands will help further our understanding of the functional interaction of agonists and GPCRs.

Prandiology of Drosophila and the CAFE assay.

Studies of feeding behavior in genetically tractable invertebrate model systems have been limited by the lack of proper methodology. We introduce the Capillary Feeder (CAFE), a method allowing precise, real-time measurement of ingestion by individual or grouped fruit flies on the scale of minutes to days. Using this technique, we conducted the first quantitative analysis of prandial behavior in Drosophila melanogaster. Our results allow the dissection of feeding into discrete bouts of ingestion, defining two separate parameters, meal volume and frequency, that can be uncoupled and thus are likely to be independently regulated. In addition, our long-term measurements show that flies can ingest as much as 1.7x their body mass over 24 h. Besides the study of appetite, the CAFE can be used to monitor oral drug delivery. As an illustration, we used the CAFE to test the effects of dietary supplementation with two compounds, paraquat and ethanol, on food ingestion and preference. Paraquat, a prooxidant widely used in stress tests, had a strong anorexigenic effect. In contrast, in a feeding preference assay, ethanol-laced food, but not ethanol by itself, acted as an attractant.

Compensatory ingestion upon dietary restriction in Drosophila melanogaster.

Dietary restriction extends the lifespan of numerous, evolutionarily diverse species. In D. melanogaster, a prominent model for research on the interaction between nutrition and longevity, dietary restriction is typically based on medium dilution, with possible compensatory ingestion commonly being neglected. Possible problems with this approach are revealed by using a method for direct monitoring of D. melanogaster feeding behavior. This demonstrates that dietary restriction elicits robust compensatory changes in food consumption. As a result, the effect of medium dilution is overestimated and, in certain cases, even fully compensated for. Our results strongly indicate that feeding behavior and nutritional composition act concertedly to determine fly lifespan. Feeding behavior thus emerges as a central element in D. melanogaster aging.