The caloric value of food intake structurally adjusts a neuronal mushroom body circuit mediating olfactory learning in Drosophila.

Associative learning enables the adaptive adjustment of behavioral decisions based on acquired, predicted outcomes. The valence of what is learned is influenced not only by the learned stimuli and their temporal relations, but also by prior experiences and internal states. In this study, we used the fruit fly Drosophila melanogaster to demonstrate that neuronal circuits involved in associative olfactory learning undergo restructuring during extended periods of low-caloric food intake. Specifically, we observed a decrease in the connections between specific dopaminergic neurons (DANs) and Kenyon cells at distinct compartments of the mushroom body. This structural synaptic plasticity was contingent upon the presence of allatostatin A receptors in specific DANs and could be mimicked optogenetically by expressing a light-activated adenylate cyclase in exactly these DANs. Importantly, we found that this rearrangement in synaptic connections influenced aversive, punishment-induced olfactory learning but did not impact appetitive, reward-based learning. Whether induced by prolonged low-caloric conditions or optogenetic manipulation of cAMP levels, this synaptic rearrangement resulted in a reduction of aversive associative learning. Consequently, the balance between positive and negative reinforcing signals shifted, diminishing the ability to learn to avoid odor cues signaling negative outcomes. These results exemplify how a neuronal circuit required for learning and memory undergoes structural plasticity dependent on prior experiences of the nutritional value of food.

The preoptic area and dorsal habenula jointly support homeostatic navigation in larval zebrafish.

Animals must maintain physiological processes within an optimal temperature range despite changes in their environment. Through behavioral assays, whole-brain functional imaging, and neural ablations, we show that larval zebrafish, an ectothermic vertebrate, achieves thermoregulation through homeostatic navigation-non-directional and directional movements toward the temperature closest to its physiological setpoint. A brain-wide circuit encompassing several brain regions enables this behavior. We identified the preoptic area of the hypothalamus (PoA) as a key brain structure in triggering non-directional reorientation when thermal conditions are worsening. This result shows an evolutionary conserved role of the PoA as principal thermoregulator of the brain also in ectotherms. We further show that the habenula (Hb)-interpeduncular nucleus (IPN) circuit retains a short-term memory of the sensory history to support the generation of coherent directed movements even in the absence of continuous sensory cues. We finally provide evidence that this circuit may not be exclusive for temperature but may convey a more abstract representation of relative valence of physiologically meaningful stimuli regardless of their specific identity to enable homeostatic navigation.

Large Language Models in Medical Education: Comparing ChatGPT- to Human-Generated Exam Questions.

PROBLEM: Creating medical exam questions is time consuming, but well-written questions can be used for test-enhanced learning, which has been shown to have a positive effect on student learning. The automated generation of high-quality questions using large language models (LLMs), such as ChatGPT, would therefore be desirable. However, there are no current studies that compare students' performance on LLM-generated questions to questions developed by humans. APPROACH: The authors compared student performance on questions generated by ChatGPT (LLM questions) with questions created by medical educators (human questions). Two sets of 25 multiple-choice questions (MCQs) were created, each with 5 answer options, 1 of which was correct. The first set of questions was written by an experienced medical educator, and the second set was created by ChatGPT 3.5 after the authors identified learning objectives and extracted some specifications from the human questions. Students answered all questions in random order in a formative paper-and-pencil test that was offered leading up to the final summative neurophysiology exam (summer 2023). For each question, students also indicated whether they thought it had been written by a human or ChatGPT. OUTCOMES: The final data set consisted of 161 participants and 46 MCQs (25 human and 21 LLM questions). There was no statistically significant difference in item difficulty between the 2 question sets, but discriminatory power was statistically significantly higher in human than LLM questions (mean = .36, standard deviation [SD] = .09 vs mean = .24, SD = .14; P = .001). On average, students identified 57% of question sources (human or LLM) correctly. NEXT STEPS: Future research should replicate the study procedure in other contexts (e.g., other medical subjects, semesters, countries, and languages). In addition, the question of whether LLMs are suitable for generating different question types, such as key feature questions, should be investigated.

Optimized design and in vivo application of optogenetically functionalized Drosophila dopamine receptors.

Neuromodulatory signaling via G protein-coupled receptors (GPCRs) plays a pivotal role in regulating neural network function and animal behavior. The recent development of optogenetic tools to induce G protein-mediated signaling provides the promise of acute and cell type-specific manipulation of neuromodulatory signals. However, designing and deploying optogenetically functionalized GPCRs (optoXRs) with accurate specificity and activity to mimic endogenous signaling in vivo remains challenging. Here we optimize the design of optoXRs by considering evolutionary conserved GPCR-G protein interactions and demonstrate the feasibility of this approach using two Drosophila Dopamine receptors (optoDopRs). These optoDopRs exhibit high signaling specificity and light sensitivity in vitro. In vivo, we show receptor and cell type-specific effects of dopaminergic signaling in various behaviors, including the ability of optoDopRs to rescue the loss of the endogenous receptors. This work demonstrates that optoXRs can enable optical control of neuromodulatory receptor-specific signaling in functional and behavioral studies.

The ecology of nutrient sensation and perception in insects.

Insects are equipped with neurological, physiological, and behavioral tools to locate potential food sources and assess their nutritional quality based on volatile and chemotactile cues. We summarize current knowledge on insect taste perception and the different modalities of reception and perception. We suggest that the neurophysiological mechanisms of reception and perception are closely linked to the species-specific ecology of different insects. Understanding these links consequently requires a multidisciplinary approach. We also highlight existing knowledge gaps, especially in terms of the exact ligands of receptors, and provide evidence for a perceptional hierarchy suggesting that insects have adapted their reception and perception to preferentially perceive nutrient stimuli that are important for their fitness.

Global change in brain state during spontaneous and forced walk in Drosophila is composed of combined activity patterns of different neuron classes.

Movement-correlated brain activity has been found across species and brain regions. Here, we used fast whole brain lightfield imaging in adult Drosophila to investigate the relationship between walk and brain-wide neuronal activity. We observed a global change in activity that tightly correlated with spontaneous bouts of walk. While imaging specific sets of excitatory, inhibitory, and neuromodulatory neurons highlighted their joint contribution, spatial heterogeneity in walk- and turning-induced activity allowed parsing unique responses from subregions and sometimes individual candidate neurons. For example, previously uncharacterized serotonergic neurons were inhibited during walk. While activity onset in some areas preceded walk onset exclusively in spontaneously walking animals, spontaneous and forced walk elicited similar activity in most brain regions. These data suggest a major contribution of walk and walk-related sensory or proprioceptive information to global activity of all major neuronal classes.

Decision making: Dopaminergic neurons for better or worse.

All animals constantly need to weigh their options based on new experiences: something initially considered bad can become better in the light of something worse. A new study now shows how flies re-evaluate between better and worse.

A dopamine-gated learning circuit underpins reproductive state-dependent odor preference in Drosophila females.

Motherhood induces a drastic, sometimes long-lasting, change in internal state and behavior in many female animals. How a change in reproductive state or the discrete event of mating modulates specific female behaviors is still incompletely understood. Using calcium imaging of the whole brain of Drosophila females, we find that mating does not induce a global change in brain activity. Instead, mating modulates the pheromone response of dopaminergic neurons innervating the fly's learning and memory center, the mushroom body (MB). Using the mating-induced increased attraction to the odor of important nutrients, polyamines, we show that disruption of the female fly's ability to smell, for instance the pheromone cVA, during mating leads to a reduction in polyamine preference for days later indicating that the odor environment at mating lastingly influences female perception and choice behavior. Moreover, dopaminergic neurons including innervation of the ß'1 compartment are sufficient to induce the lasting behavioral increase in polyamine preference. We further show that MB output neurons (MBON) of the ß'1 compartment are activated by pheromone odor and their activity during mating bidirectionally modulates preference behavior in mated and virgin females. Their activity is not required, however, for the expression of polyamine attraction. Instead, inhibition of another type of MBON innervating the ß'2 compartment enables expression of high odor attraction. In addition, the response of a lateral horn (LH) neuron, AD1b2, which output is required for the expression of polyamine attraction, shows a modulated polyamine response after mating. Taken together, our data in the fly suggests that mating-related sensory experience regulates female odor perception and expression of choice behavior through a dopamine-gated learning circuit.

A role for glia in cellular and systemic metabolism: insights from the fly.

Excitability and synaptic transmission make neurons high-energy consumers. However, neurons do not store carbohydrates or lipids. Instead, they need support cells to fuel their metabolic demands. This role is assumed by glia, both in vertebrates and invertebrates. Many questions remain regarding the coupling between neuronal activity and energy demand on the one hand, and nutrient supply by glia on the other hand. Here, we review recent advances showing that fly glia, similar to their role in vertebrates, fuel neurons in times of high energetic demand, such as during memory formation and long-term storage. Vertebrate glia also play a role in the modulation of neurons, their communication, and behavior, including food search and feeding. We discuss recent literature pointing to similar roles of fly glia in behavior and metabolism.

Action selection: Neuropeptidergic gates of behavior.

Nervous systems continuously receive environmental signals with distinct behavioral meanings. To process ambiguous sensory inputs, neural circuits rely on hubs with compartmentalized synaptic structures. A new study has revealed how, in Drosophila larvae, this architecture with the local release of neuropeptides enables the control of flexible and context-dependent behavioral outcomes.

Preparing Adult Drosophila melanogaster for Whole Brain Imaging during Behavior and Stimuli Responses.

We present a method developed specifically to image the whole Drosophila brain during ongoing behavior such as walking. Head fixation and dissection are optimized to minimize their impact on behavior. This is first achieved by using a holder that minimizes movement hindrances. The back of the fly's head is glued to this holder at an angle that allows optical access to the whole brain while retaining the fly's ability to walk, groom, smell, taste and see. The back of the head is dissected to remove tissues in the optical path and muscles responsible for head movement artefacts. The fly brain can subsequently be imaged to record brain activity, for instance using calcium or voltage indicators, during specific behaviors such as walking or grooming, and in response to different stimuli. Once the challenging dissection, which requires considerable practice, has been mastered, this technique allows to record rich data sets relating whole brain activity to behavior and stimulus responses.

Flies Avoid Current Atmospheric CO2 Concentrations.

CO2 differs from most other odors by being ubiquitously present in the air animals inhale. CO2 levels of the atmosphere, however, are subject to change. Depending on the landscape, temperature, and time of the year, CO2 levels can change even on shortest time scales. In addition, since the 18th century the CO2 baseline keeps increasing due to the intensive fossil fuel usage. However, we do not know whether this change is significant for animals, and if yes whether and how animals adapt to this change. Most insects possess olfactory receptors to detect the gaseous molecule, and CO2 is one of the key odorants for insects such as the vinegar fly Drosophila melanogaster to find food sources and to warn con-specifics. So far, CO2 and its sensory system have been studied in the context of rotting fruit and other CO2-emitting sources to investigate flies' response to significantly elevated levels of CO2. However, it has not been addressed whether flies detect and potentially react to atmospheric levels of CO2. By using behavioral experiments, here we show that flies can detect atmospheric CO2 concentrations and, if given the choice, prefer air with sub-atmospheric levels of the molecule. Blocking the synaptic release from CO2 receptor neurons abolishes this choice. Based on electrophysiological recordings, we hypothesize that CO2 receptors, similar to ambient temperature receptors, actively sample environmental CO2 concentrations close to atmospheric levels. Based on recent findings and our data, we hypothesize that Gr-dependent CO2 receptors do not primarily serve as a cue detector to find food sources or avoid danger, instead they function as sensors for preferred environmental conditions.

Dopamine modulation of sensory processing and adaptive behavior in flies.

Behavioral flexibility for appropriate action selection is an advantage when animals are faced with decisions that will determine their survival or death. In order to arrive at the right decision, animals evaluate information from their external environment, internal state, and past experiences. How these different signals are integrated and modulated in the brain, and how context- and state-dependent behavioral decisions are controlled are poorly understood questions. Studying the molecules that help convey and integrate such information in neural circuits is an important way to approach these questions. Many years of work in different model organisms have shown that dopamine is a critical neuromodulator for (reward based) associative learning. However, recent findings in vertebrates and invertebrates have demonstrated the complexity and heterogeneity of dopaminergic neuron populations and their functional implications in many adaptive behaviors important for survival. For example, dopaminergic neurons can integrate external sensory information, internal and behavioral states, and learned experience in the decision making circuitry. Several recent advances in methodologies and the availability of a synaptic level connectome of the whole-brain circuitry of Drosophila melanogaster make the fly an attractive system to study the roles of dopamine in decision making and state-dependent behavior. In particular, a learning and memory center-the mushroom body-is richly innervated by dopaminergic neurons that enable it to integrate multi-modal information according to state and context, and to modulate decision-making and behavior.

Immune Receptor Signaling and the Mushroom Body Mediate Post-ingestion Pathogen Avoidance.

In spite of the positive effects of bacteria on health, certain species are harmful, and therefore, animals must weigh nutritional benefits against negative post-ingestion consequences and adapt their behavior accordingly. Here, we use Drosophila to unravel how the immune system communicates with the brain, enabling avoidance of harmful foods. Using two different known fly pathogens, mildly pathogenic Erwinia carotovora (Ecc15) and highly virulent Pseudomonas entomophila (Pe), we analyzed preference behavior in naive flies and after ingestion of either of these pathogens. Although survival assays confirmed the harmful effect of pathogen ingestion, naive flies preferred the odor of either pathogen to air and also to harmless mutant bacteria, suggesting that flies are not innately repelled by these microbes. By contrast, feeding assays showed that, when given a choice between pathogenic and harmless bacteria, flies-after an initial period of indifference-shifted to a preference for the harmless strain, a behavior that lasted for several hours. Flies lacking synaptic output of the mushroom body (MB), the fly's brain center for associative memory formation, lost the ability to distinguish between pathogenic and harmless bacteria, suggesting this to be an adaptive behavior. Interestingly, this behavior relied on the immune receptors PGRP-LC and -LE and their presence in octopaminergic neurons. We postulate a model wherein pathogen ingestion triggers PGRP signaling in octopaminergic neurons, which in turn relay the information about the harmful food source directly or indirectly to the MB, where an appropriate behavioral output is generated.

Sensory Evolution: Making Sense of the Noni Scent.

A recent study identifies the neuronal and molecular underpinnings of a key ecological difference between two Drosophila species using a remarkable genetic toolbox for a non-model species.

Valence and State-Dependent Population Coding in Dopaminergic Neurons in the Fly Mushroom Body.

Neuromodulation permits flexibility of synapses, neural circuits, and ultimately behavior. One neuromodulator, dopamine, has been studied extensively in its role as a reward signal during learning and memory across animal species. Newer evidence suggests that dopaminergic neurons (DANs) can modulate sensory perception acutely, thereby allowing an animal to adapt its behavior and decision making to its internal and behavioral state. In addition, some data indicate that DANs are not homogeneous but rather convey different types of information as a heterogeneous population. We have investigated DAN population activity and how it could encode relevant information about sensory stimuli and state by taking advantage of the confined anatomy of DANs innervating the mushroom body (MB) of the fly Drosophila melanogaster. Using in vivo calcium imaging and a custom 3D image registration method, we found that the activity of the population of MB DANs encodes innate valence information of an odor or taste as well as the physiological state of the animal. Furthermore, DAN population activity is strongly correlated with movement, consistent with a role of dopamine in conveying behavioral state to the MB. Altogether, our data and analysis suggest that DAN population activities encode innate odor and taste valence, movement, and physiological state in a MB-compartment-specific manner. We propose that dopamine shapes innate perception through combinatorial population coding of sensory valence, physiological, and behavioral context.

Studying complex brain dynamics using Drosophila.

The field has successfully used Drosophila genetic tools to identify neurons and sub-circuits important for specific functions. However, for an organism with complex and changing internal states to succeed in a complex and changing natural environment, many neurons and circuits need to interact dynamically. Drosophila's many advantages, combined with new imaging tools, offer unique opportunities to study how the brain functions as a complex dynamical system. We give an overview of complex activity patterns and how they can be observed, as well as modeling strategies, adding proof of principle in some cases.

A Neural Circuit Arbitrates between Persistence and Withdrawal in Hungry Drosophila.

In pursuit of food, hungry animals mobilize significant energy resources and overcome exhaustion and fear. How need and motivation control the decision to continue or change behavior is not understood. Using a single fly treadmill, we show that hungry flies persistently track a food odor and increase their effort over repeated trials in the absence of reward suggesting that need dominates negative experience. We further show that odor tracking is regulated by two mushroom body output neurons (MBONs) connecting the MB to the lateral horn. These MBONs, together with dopaminergic neurons and Dop1R2 signaling, control behavioral persistence. Conversely, an octopaminergic neuron, VPM4, which directly innervates one of the MBONs, acts as a brake on odor tracking by connecting feeding and olfaction. Together, our data suggest a function for the MB in internal state-dependent expression of behavior that can be suppressed by external inputs conveying a competing behavioral drive.

Flies spring a surprise.

A combination of genetic, anatomical and physiological techniques has revealed that the lateral horn, a region of the brain involved in olfaction in flies, has many more types of neurons than expected.