Edgar Buhl

Drosophila Ionotropic Receptor 25a mediates circadian clock resetting by temperature

Circadian clocks are endogenous timers adjusting behaviour and physiology with the solar day. Synchronized circadian clocks improve fitness and are crucial for our physical and mental well-being. Visual and non-visual photoreceptors are responsible for synchronizing circadian clocks to light, but clock-resetting is also achieved by alternating day and night temperatures with only 2–4 °C difference. This temperature sensitivity is remarkable considering that the circadian clock period (~24 h) is largely independent of surrounding ambient temperatures. Here we show that Drosophila Ionotropic Receptor 25a (IR25a) is required for behavioural synchronization to low-amplitude temperature cycles. This channel is expressed in sensory neurons of internal stretch receptors previously implicated in temperature synchronization of the circadian clock. IR25a is required for temperature-synchronized clock protein oscillations in subsets of central clock neurons. Extracellular leg nerve recordings reveal temperature- and IR25a-dependent sensory responses, and IR25a misexpression confers temperature-dependent firing of heterologous neurons. We propose that IR25a is part of an input pathway to the circadian clock that detects small temperature differences. This pathway operates in the absence of known ‘hot’ and ‘cold’ sensors in the Drosophila antenna, revealing the existence of novel periphery-to-brain temperature signalling channels.

A muscarinic cholinergic mechanism underlies activation of the central pattern generator for locust flight

SUMMARY A central question in behavioural control is how central pattern generators (CPGs) for locomotion are activated. This paper disputes the key role generally accredited to octopamine in activating the CPG for insect flight. In deafferented locusts, fictive flight was initiated by bath application of the muscarinic agonist pilocarpine, the acetylcholine analogue carbachol, and the acetylcholinesterase blocker eserine, but not by nicotine. Furthermore, in addition to octopamine, various other amines including dopamine, tyramine and histamine all induced fictive flight, but not serotonin or the amine-precursor amino acid tyrosine. However, flight initiation was not reversibly blocked by aminergic antagonists, and was still readily elicited by both natural stimulation (wind) and pilocarpine in reserpinized, amine-depleted locusts. By contrast, the muscarinic antagonists atropine and scopolamine reversibly blocked flight initiated by wind, cholinergic agonists, octopamine, and by selective stimulation of a flight-initiating interneurone (TCG). The short delay from TCG stimulation to flight onset suggests that TCG acts directly on the flight CPG, and accordingly that TCG, or its follower cell within the flight generating circuit, is cholinergic. We conclude that acetylcholine acting via muscarinic receptors is the key neurotransmitter in the mechanism underlying the natural activation of the locust flight CPG. Amines are not essential for this, but must be considered as potential neuromodulators for facilitating flight release and tuning the motor pattern. We speculate that muscarinic activation coupled to aminergic facilitation may be a general feature of behavioural control in insects for ensuring conditional recruitment of individual motor programs in accordance with momentary adaptive requirements.

Can Simple Rules Control Development of a Pioneer Vertebrate Neuronal Network Generating Behavior

How do the pioneer networks in the axial core of the vertebrate nervous system first develop? Fundamental to understanding any full-scale neuronal network is knowledge of the constituent neurons, their properties, synaptic interconnections, and normal activity. Our novel strategy uses basic developmental rules to generate model networks that retain individual neuron and synapse resolution and are capable of reproducing correct, whole animal responses. We apply our developmental strategy to young Xenopus tadpoles, whose brainstem and spinal cord share a core vertebrate plan, but at a tractable complexity. Following detailed anatomical and physiological measurements to complete a descriptive library of each type of spinal neuron, we build models of their axon growth controlled by simple chemical gradients and physical barriers. By adding dendrites and allowing probabilistic formation of synaptic connections, we reconstruct network connectivity among up to 2000 neurons. When the resulting “network” is populated by model neurons and synapses, with properties based on physiology, it can respond to sensory stimulation by mimicking tadpole swimming behavior. This functioning model represents the most complete reconstruction of a vertebrate neuronal network that can reproduce the complex, rhythmic behavior of a whole animal. The findings validate our novel developmental strategy for generating realistic networks with individual neuron- and synapse-level resolution. We use it to demonstrate how early functional neuronal connectivity and behavior may in life result from simple developmental “rules,” which lay out a scaffold for the vertebrate CNS without specific neuron-to-neuron recognition.

A Neural Network Underlying Circadian Entrainment and Photoperiodic Adjustment of Sleep and Activity in Drosophila

A sensitivity of the circadian clock to light/dark cycles ensures that biological rhythms maintain optimal phase relationships with the external day. In animals, the circadian clock neuron network (CCNN) driving sleep/activity rhythms receives light input from multiple photoreceptors, but how these photoreceptors modulate CCNN components is not well understood. Here we show that the Hofbauer-Buchner eyelets differentially modulate two classes of ventral lateral neurons (LNvs) within the Drosophila CCNN. The eyelets antagonize Cryptochrome (CRY)- and compound-eye-based photoreception in the large LNvs while synergizing CRY-mediated photoreception in the small LNvs. Furthermore, we show that the large LNvs interact with subsets of “evening cells” to adjust the timing of the evening peak of activity in a day length-dependent manner. Our work identifies a peptidergic connection between the large LNvs and a group of evening cells that is critical for the seasonal adjustment of circadian rhythms. SIGNIFICANCE STATEMENT In animals, circadian clocks have evolved to orchestrate the timing of behavior and metabolism. Consistent timing requires the entrainment these clocks to the solar day, a process that is critical for an organisms health. Light cycles are the most important external cue for the entrainment of circadian clocks, and the circadian system uses multiple photoreceptors to link timekeeping to the light/dark cycle. How light information from these photorecptors is integrated into the circadian clock neuron network to support entrainment is not understood. Our results establish that input from the HB eyelets differentially impacts the physiology of neuronal subgroups. This input pathway, together with input from the compound eyes, precisely times the activity of flies under long summer days. Our results provide a mechanistic model of light transduction and integration into the circadian system, identifying new and unexpected network motifs within the circadian clock neuron network.

Female crickets are driven to fight by the male courting and calling songs

Crickets have traditional sex roles, where males compete aggressively for access to selective polyandrous females. However, in a laboratory experiment, we found that normally nonaggressive female Gryllus campestris fought each other vigorously in the presence of a courting male, resulting in a dominant female that gained a greater probability of receiving the spermatophore. Female–female fights included the same series of characteristic actions known from male–male fights, which demonstrates that the females can perform the full repertoire of agonistic actions except for the production of the aggressive rival song. Since females remained nonaggressive towards each other in the vicinity of a muted male, but were induced to fight each other in the complete absence of a male by the auditory experience of the courtship song, this song is both sufficient and necessary to induce female competition for males. Calling song was as effective as courtship song at inducing female aggression, whereas rival song was least effective. We therefore speculate that the calling and courtship songs may signal a males resource value.

The role of a trigeminal sensory nucleus in the initiation of locomotion

Key points  •  Rhythmic activity is a feature of many regions of the CNS, but surprisingly, no precise pathway for the initiation of locomotion is yet known for any vertebrate. •  Using a well‐proven, simple vertebrate system, the hatchling Xenopus tadpole, we report here our discovery of a detailed neuron‐by‐neuron pathway for initiating locomotor activity on one side of the CNS. •  We describe a small population of brainstem neurons (trigeminal interneurons) that are directly excited by trigeminal sensory neurons when the head skin is touched. •  These neurons amplify brief sensory signals and relay excitation to an electrically coupled population of hindbrain reticulospinal neurons (descending interneurons), whose firing initiates swimming locomotion. •  We believe that our discovery of this primitive, direct pathway, which appears simpler than initiation pathways so far defined in invertebrates, is of evolutionary interest and raises the important possibility of equivalent pathways in more complex vertebrates including mammals.

Quasimodo mediates daily and acute light effects on Drosophila clock neuron excitability

Significance Most organisms developed a circadian clock to adapt their behavior to daily changes of light and temperature. The molecular clock is remarkably conserved across species with much of our current understanding coming from Drosophila studies. To generate circadian behavior, appropriate levels of neuronal electrical activity are crucial, but the regulators of this activity have remained largely elusive. Here we identify three membrane proteins that interact to set the clock neurons to “day” or “night,” forming a light-input pathway to the circadian clock. The membrane-anchored extracellular protein Quasimodo affects both the daily changes in physiological properties and light responses of brain clock neurons, possibly acting upstream of the potassium channel Shaw and the Na+, K+, Cl− ion transporter NKCC. We have characterized a light-input pathway regulating Drosophila clock neuron excitability. The molecular clock drives rhythmic electrical excitability of clock neurons, and we show that the recently discovered light-input factor Quasimodo (Qsm) regulates this variation, presumably via an Na+, K+, Cl− cotransporter (NKCC) and the Shaw K+ channel (dKV3.1). Because of light-dependent degradation of the clock protein Timeless (Tim), constant illumination (LL) leads to a breakdown of molecular and behavioral rhythms. Both overexpression (OX) and knockdown (RNAi) of qsm, NKCC, or Shaw led to robust LL rhythmicity. Whole-cell recordings of the large ventral lateral neurons (l-LNv) showed that altering Qsm levels reduced the daily variation in neuronal activity: qsmOX led to a constitutive less active, night-like state, and qsmRNAi led to a more active, day-like state. Qsm also affected daily changes in K+ currents and the GABA reversal potential, suggesting a role in modifying membrane currents and GABA responses in a daily fashion, potentially modulating light arousal and input to the clock. When directly challenged with blue light, wild-type l-LNvs responded with increased firing at night and no net response during the day, whereas altering Qsm, NKKC, or Shaw levels abolished these day/night differences. Finally, coexpression of ShawOX and NKCCRNAi in a qsm mutant background restored LL-induced behavioral arrhythmicity and wild-type neuronal activity patterns, suggesting that the three genes operate in the same pathway. We propose that Qsm affects both daily and acute light effects in l-LNvs probably acting on Shaw and NKCC.

Sensory initiation of a co‐ordinated motor response: synaptic excitation underlying simple decision‐making

Deciding whether or how to initiate a motor response to a stimulus can be surprisingly slow and the underlying processes are not well understood. The neuronal circuitry that allows frog tadpoles to swim in response to touch is well characterized and includes excitatory reticulospinal neurons that drive swim circuit neurons. Build‐up of excitation to reticulospinal neurons is the key decision‐making step for swimming. Asymmetry in this build‐up between the two sides allows bilateral initiation at the same time as avoiding inappropriate co‐activation of motor antagonists. Following stronger stimuli, reticulospinal neurons are excited through a trigeminal nucleus pathway and swimming starts first on the stimulated side. If this pathway fails or is lesioned, swimming starts later on the unstimulated side. The mechanisms underlying initiation of a simple tadpole motor response may share similarities with more complex decisions in other animals, including humans.

A lineage-related reciprocal inhibition circuitry for sensory-motor action selection

The insect central complex and vertebrate basal ganglia are forebrain centres involved in selection and maintenance of behavioural actions. However, little is known about the formation of the underlying circuits, or how they integrate sensory information for motor actions. Here, we show that paired embryonic neuroblasts generate central complex ring neurons that mediate sensory-motor transformation and action selection in Drosophila. Lineage analysis resolves four ring neuron subtypes, R1-R4, that form GABAergic inhibition circuitry among inhibitory sister cells. Genetic manipulations, together with functional imaging, demonstrate subtype-specific R neurons mediate the selection and maintenance of behavioural activity. A computational model substantiates genetic and behavioural observations suggesting that R neuron circuitry functions as salience detector using competitive inhibition to amplify, maintain or switch between activity states. The resultant gating mechanism translates facilitation, inhibition and disinhibition of behavioural activity as R neuron functions into selection of motor actions and their organisation into action sequences.

Drosophila PINK1 and parkin loss-of-function mutants display a range of non-motor Parkinson's disease phenotypes

Parkinsons disease (PD) is more commonly associated with its motor symptoms and the related degeneration of dopamine (DA) neurons. However, it is becoming increasingly clear that PD patients also display a wide range of non-motor symptoms, including memory deficits and disruptions of their sleep-wake cycles. These have a large impact on their quality of life, and often precede the onset of motor symptoms, but their etiology is poorly understood. The fruit fly Drosophila has already been successfully used to model PD, and has been used extensively to study relevant non-motor behaviours in other contexts, but little attention has yet been paid to modelling non-motor symptoms of PD in this genetically tractable organism. We examined memory performance and circadian rhythms in flies with loss-of-function mutations in two PD genes: PINK1 and parkin. We found learning and memory abnormalities in both mutant genotypes, as well as a weakening of circadian rhythms that is underpinned by electrophysiological changes in clock neurons. Our study paves the way for further work that may help us understand the mechanisms underlying these neglected aspects of PD, thus identifying new targets for treatments to address these non-motor problems specifically and perhaps even to halt disease progression in its prodromal phase.