InteractiveFly: GeneBrief

Histamine-gated chloride channel subunit 1: Biological Overview | References

In Drosophila, the clock that controls rest-activity rhythms synchronizes with light-dark cycles through either the blue-light sensitive cryptochrome (Cry) located in most clock neurons, or rhodopsin-expressing histaminergic photoreceptors. This study shows that, in the absence of Cry, each of the two histamine receptors Ort and HisCl1 contribute to entrain the clock whereas no entrainment occurs in the absence of the two receptors. In contrast to Ort, HisCl1 does not restore entrainment when expressed in the optic lobe interneurons. Indeed, HisCl1 is expressed in wild-type photoreceptors and entrainment is strongly impaired in flies with photoreceptors mutant for HisCl1. Rescuing HisCl1 expression in the Rh6-expressing photoreceptors restores entrainment but it does not in other photoreceptors, which send histaminergic inputs to Rh6-expressing photoreceptors. These results thus show that Rh6-expressing neurons contribute to circadian entrainment as both photoreceptors and interneurons, recalling the dual function of melanopsin-expressing ganglion cells in the mammalian retina (Alejevski, 2019).

The Drosophila sleep-wake rhythms are controlled by a brain circadian clock that includes about 150 clock neurons. Light synchronizes the clock neuronal network through cell-autonomous and non-cell-autonomous light input pathways. Cry is a blue-light sensitive photoreceptor protein that is expressed in most clock neurons. In the absence of Cry, flies do not phase-shift their behavioral rhythms in response to a short light pulse but still synchronize to light-dark (LD) cycles. Only flies devoid of both Cry and rhodopsin-expressing photoreceptors fail to entrain to LD cycles. Six different rhodopsins (Rhs) have been characterized in the Drosophila photoreceptive structures, which include the compound eye, the Hofbauer-Buchner (H-B) eyelet, and ocelli. The compound eye strongly contributes to circadian photoreception, whereas a modest contribution appears to be brought by the H-B eyelet and the ocelli. A circadian function has been recently associated with the yet poorly characterized rhodopsin 7, although its exact contribution and localization in the brain and/or the eye remains controversial. In addition to entrainment, the visual system controls other features of the clock neuron network by conveying light information to either promote or inhibit the behavioral output of specific clock neuron subsets (Alejevski, 2019).

The compound eye includes about 800-unit eyes (ommatidia), each of which contains eight photoreceptors. The six Rh1-expressing outer photoreceptors (R1-6) are involved in motion detection and project to the lamina neuropile of the optic lobe. The two inner photoreceptors (R7-8) are important for color detection and project to the medulla. They express four different rhodopsins and thus define two types of ommatidia: 'pale' (p) ommatidia (30%) include a Rh3-expressing R7 and a Rh5-expressing R8, whereas 'yellow' (y) ommatidia (70%) include a Rh4-expressing R7 and a Rh6-expressing R8. Each extra-retinal H-B eyelet contains four Rh6-expressing photoreceptors that project to the accessory medulla, in the vicinity of key pacemaker neurons, the ventral lateral neurons (LNvs) that produce the pigment-dispersing factor (PDF) neuropeptide9,20-24. Each of the three ocelli contains about 80 photoreceptors that express Rh225. The Drosophila rhodopsins cover a wide range of wavelengths from 300 nm to 600 nm18,19, with only Rh1 and Rh6 being sensitive to red light (Alejevski, 2019).

Rhodopsin-dependent circadian entrainment involves two downstream signaling pathways, the canonical one that relies on the phospholipase C encoded by the no receptor potential A gene (norpA)2 or an unknown pathway that does not contribute in very low light levels. All but Rh2- and Rh5- expressing photoreceptors support synchronization in very low light, and at least Rh1, Rh5, and Rh6 can signal through the NorpA-independent pathway. Photoreceptors of the compound eye are histaminergic but the H-B eyelet expresses both histamine and acetylcholine. Although the two neurotransmitters might contribute to circadian entrainment, flies devoid of Cry and histidine decarboxylase do not synchronize their rest-activity rhythms with LD cycles. This suggests that besides Cry, there is no histamine-independent pathway to entrain the clock (Alejevski, 2019).

Two genes encoding histamine-gated chloride channels, ora transientless (ort) and Histamine-gated chloride channel subunit 1 (HisCl1), have been identified in Drosophila. The ort-null mutants are visually blind and their electroretinograms have no ON and OFF transients. In contrast, HisCl1 mutants show increased OFF transients, whereas slower responses were observed in the postsynaptic laminar monopolar cells. Based on transcriptional reporters, ort expression in the optic lobes was observed in neurons of both the lamina and medulla/lobula neuropils. Based on reporter gene expression, HisCl1 was localized in glial cells of the lamina. However, recent work reported expression in photoreceptors, in particular in the R7 and R8 inner photoreceptor subtypes. Indeed, Ort and HisCl1 support color opponency between the two subtypes of 'inner' photoreceptors, the ultraviolet (UV)-sensitive R7 and non-UV-sensitive R8, with HisCl1 and Ort mediating direct and indirect inhibition, respectively. The histaminergic pathways that are involved in circadian entrainment are unknown and are the subject of the present study. The results show that both Ort and HisCl1 define two different pathways for circadian entrainment. Whereas Ort contributes through its expression in the interneurons of the optic lobe, HisCl1 mostly contributes through its expression in the Rh6-expressing retinal photoreceptors. The work thus reveals that Rh6-expressing neurons contribute to light-mediated entrainment as both photoreceptors and interneurons (Alejevski, 2019).

This work reveals that the Cryptochrome-independent entrainment of rest-activity rhythms relies on distinct pathways that are contributed by the two histamine receptors Ort and HisCl1. Whereas Ort mediates circadian entrainment through the optic lobe interneurons that are involved in visual functions, HisCl1 defines a new photoreceptive pathway through Rh6-expressing photoreceptors. Although both receptors mediate synchronization with a shifted LD cycle, it seems likely that the two pathways will show differences in specific light conditions. It was not possible to rescue Ort function with HisCl1 expression in the ort-expressing cells, whereas the Ort could replace HisCl1 in Rh6 photoreceptors. It is possible that HisCl1 has a lower affinity for histamine with Rh6 cells receiving more neurotransmitter than optic lobe interneurons. Alternatively, interneurons could sufficiently differ from photoreceptors for their physiology or specific receptor-interacting protein content, preventing HisCl1 from working efficiently. HisCl1 downregulation in Rh6 cells slows down synchronization and flies with HisCl1134 mutant eyes synchronize very poorly with advanced LD cycles, and fail to synchronize with delays. It cannot be excluded that non-photoreceptor cells contribute to HisCl1-dependent entrainment but other pathways appear to have a modest contribution if any (Alejevski, 2019).

HisCl1 is expressed in the H-B eyelet, which could thus contribute to this synchronization pathway. However, the cell-killing experiments indicate that H-B eyelet is not required for HisCl1-mediated synchronization through Rh6 cells. In the recently described color opponency mechanism, retinal R7 cells inhibit R8 and vice versa through HisCl1 expression in the photoreceptors (Schnaitmann, 2018). It is supposed that HisCl1-dependent clock synchronization is also mediated by the hyperpolarization of Rh6-expressing cells. How this hyperpolarization interacts with the light-induced depolarization in Rh6 photoreceptors to result in a synchronization message to the clock neurons remains to be understood. Since only Rh6-expressing R8 and not the other inner photoreceptors contribute to this circadian photoreception pathway, Rh6 cells might have specific connections with downstream interneurons. Such specificity has been described for color vision where each of the four inner photoreceptor subtypes connects to a different type of TmY interneuron in the Medulla. This study shows that HisCl1 expression in Rh6 cells supports synchronization with red light, in the absence of Rh1, indicating that an intra-Rh6-photoreceptor circuit is sufficient. This indicates that Rh6-expressing R8 photoreceptors play a dual photoreceptor/interneuron role in this pathway (Model for the retinal input pathways to the brain clock). Whether the same individual cells have the two roles is unknown, although the HisCl1-dependent color opponency mechanism suggests that it could be the case. It is also unclear whether all Rh6-expressing R8 photoreceptors or only a fraction of them contribute to circadian synchronization. The results imply that, in addition to histaminergic neurotransmission, Rh6-expressing photoreceptors can talk to downstream interneurons through histamine-independent neurotransmission. A recent transcriptomics study indeed revealed the expression of cholinergic markers in R7 and R8 cells, supporting cholinergic transmission in the inner photoreceptors, in addition to histaminergic transmission (Alejevski, 2019).

The data indicate that histaminergic inputs from both outer and inner photoreceptors converge to Rh6 cells to contribute to circadian entrainment. It is possible that some of these inputs rely on Rh7, which seems to be expressed in Rh6-expressing photoreceptors, according to transcriptional reporter data. Putative connections between photoreceptors have been described in Drosophila and other insects. How R1-6 photoreceptors might be connected to Rh6-expressing R8 cells remains difficult to understand, but a few putative contacts between presynaptic outer cells and postsynaptic inner cells have been observed in Musca. The intra-retinal functional connectivity that this study reports in Drosophila is reminiscent to the circuit logic of circadian entrainment in the mammalian retina, where intrinsically photoreceptive retinal ganglion cells express the melanopsin photopigment in addition to receiving inputs from rods and cones. Interestingly, melanopsin appears to share light-sensing properties with the rhabdomeric photoreceptors of invertebrates. It has been shown that the mammalian circadian clock can synchronize with day-night cycles by tracking light color changes in addition to light intensity changes. It will be interesting to investigate the possible contribution of the dual function of Rh6-expressing photoreceptors to integrating different color cues into the retinal information that is sent to the clock (Alejevski, 2019).

Color Processing in the Early Visual System of Drosophila

Color vision is encoded by color-opponent neurons that are excited at one wavelength and inhibited at another. This study examined the circuit implementation of color-opponent processing in the Drosophila visual system by combining two-photon calcium imaging with genetic dissection of visual circuits. Color-opponent processing of UVshort/blue and UVlong/green is already implemented in R7/R8 inner photoreceptor terminals of "pale" and "yellow" ommatidia, respectively. R7 and R8 photoreceptors of the same type of ommatidia mutually inhibit each other directly via HisCl1 histamine receptors (see the Graphical Abstract) and receive additional feedback inhibition that requires the second histamine receptor Ort. Color-opponent processing at the first visual synapse represents an unexpected commonality between Drosophila and vertebrates; however, the differences in the molecular and cellular implementation suggest that the same principles evolved independently (Schnaitmann. 2018).

Color vision enables animals to distinguish spectral stimuli independent of their relative intensities and provides an extra dimension to vision that facilitates discrimination tasks and intra-specific communication. In pollinators such as honeybees, color vision plays a crucial role in flower recognition and, thus, has both ecological and economic importance. Color vision requires possession of at least two photoreceptor types with different spectral sensitivities and the ability to compare their outputs. Antagonistic interactions between different channels are a hallmark of sensory processing that enhances stimulus contrast and maximizes information transfer. In color vision, this opponency between spectral channels solves the critical shortcoming that any single photoreceptor cannot distinguish between changes in brightness and spectral information (Schnaitmann. 2018).

Much knowledge of color-opponent processing is based on vertebrates. In humans and other trichromatic primates, the signals of short (S), middle (M), and long (L) wavelength-sensitive cone photoreceptors are combined antagonistically to create two spectrally opponent pathways. In the first pathway, L and M cone signals mutually inhibit each other. In the second pathway S cone signals and the summed signals of L and M cones mutually inhibit each other. These color opponencies correspond with the red-green and blue-yellow opponent axes of human and macaque color perception and are implemented at the first visual synapse. Different types of cones converge onto horizontal cells, and the latter establish reciprocal sign-inverting synapses with cone terminals. This circuit adapts photoreceptor output to the intensity of ambient light, enhances achromatic and chromatic image contrast, and renders the terminals of vertebrate cones color-opponent. Furthermore, it provides the basis for the center-surround organization of cone terminals and bipolar cells. However, the exact biophysical mechanisms that underlie this critical processing stage in the retina are still unresolved (Schnaitmann. 2018).

Compared with the vertebrate retina, less is known about color-opponent processing in insects. Color-opponent neurons have been recorded in a few species, but the lack of genetic amenability prohibited identification of the underlying circuits and synaptic mechanisms. Recent studies of sensory processing in Drosophila suggest that such insights may be revealed in this model organism; however, color-opponent neurons have not been previously identified. Such neurons are expected because fruit flies exhibit a multitude of wavelength-dependent behaviors, including phototaxis, spectral preference, and color memory. Moreover, Drosophila is a classic model for studies of visual system development and connectivity and offers almost unlimited genetic amenability. In the Drosophila eye, each ommatidium houses six outer photoreceptors, R1-R6, and a pair of superimposed inner photoreceptors, R7/R8 that release histamine as neurotransmitter. The broadband-sensitive R1-R6 photoreceptors express rhodopsin1 (rh1), project to the lamina, and provide the major input to the motion vision system. Although not required for color vision, R1-R6 photoreceptors were recently shown to also contribute to it. In R7/R8 photoreceptor pairs, precise genetic control of rhodopsin expression determines the two major types of ommatidia, 'pale' (p) and 'yellow' (y), that are stochastically distributed over the main part of the eye. R7p and R8p photoreceptors express rh3 with maximum sensitivity in the short-UV and rh5 with maximum sensitivity in the blue spectral range, respectively. R7y and R8y photoreceptors express rh4 with maximum sensitivity in the long-UV and rh6 with maximum sensitivity in the green spectral range, respectively. These four types of inner photoreceptors provide the major input to the color vision system in the medulla, to which they project without making chemical synapses in the lamina (Schnaitmann. 2018).

Direct investigation of color-opponent processing using electrophysiological recordings has so far proved elusive by the technical difficulties associated with the stacked arrangement of the R7 and R8 cells and their postsynaptic partners in Drosophila. However, behavioral studies on Lucilia and Drosophila suggest that flies compare the signals of R7 and R8 photoreceptors of the same type of ommatidia (p and y) to distinguish color. Intracellular recordings from distal segments of photoreceptors in large dipteran flies and most other insect species revealed no sign of spectral inhibition; therefore, this comparison is thought to be implemented downstream of Drosophila inner photoreceptors. Candidate color-opponent neurons postsynaptic to inner photoreceptors have been revealed in behavioral, genetic, and anatomical studies in Drosophila and include Tm9, Tm20, Tm5a/b/c, and certain TmY cells. However, these studies do not exclude the possibility of color-opponent processing at the level of photoreceptors. For example, intracellular recordings in a few butterfly species revealed excitatory and inhibitory response components in particular photoreceptors. Finally, the responses of photoreceptor terminals have not been recorded in any insect species so far. This leaves open the question of whether local inhibitory circuit mechanisms render photoreceptor output color-opponent and, if so, how these mechanisms compare with processing in the vertebrate retina (Schnaitmann. 2018).

This study reports physiological recordings from inner photoreceptor terminals of Drosophila, combining two-photon calcium imaging, spectral stimulation, and use of the fluorescent genetically encoded calcium reporter Twitch-2C. Presynaptic UVshort/blue and UVlong/green color opponencies are evident in the terminals of the inner photoreceptors R7/R8 of p and y ommatidia. Genetic dissection of the peripheral visual circuits enabled identification of the photoreceptor interactions underlying color-opponent processing: R7 and R8 photoreceptors of the same type of ommatidia mutually inhibit each other at the level of their presynaptic terminals, whereas R1-R6 do not contribute to spectrally opponent processing in R7/R8 photoreceptor terminals. Two concurrent circuit mechanisms that involve the distinct histamine receptors HisCl1 and Ort mediate this processing. Direct inhibitory synaptic interactions between the terminals of R7/R8 pairs are mediated by HisCl1, and feedback inhibition with similar spectral tuning requires expression of Ort. These results illustrate that the Drosophila visual system subtracts different spectral channels at the first synapse, reminiscent of processing in the vertebrate retina, albeit by entirely different synaptic and cellular mechanisms (Schnaitmann. 2018).

Based on the spectral sensitivities of the Rhodopsins expressed in the four types of inner photoreceptors in Drosophila, R7 and R8 cells have been implicated in color vision for decades. Recent studies corroborated this view and provided further insights into the neural underpinnings of Drosophila color vision. However, because of the lack of physiological recordings from neurons in the color pathway, it is still unknown how color information is processed in Drosophila at the cellular and circuit level. This study reports an optophysiological approach that enables the analysis of neural responses to spectral stimuli in Drosophila. Based on this approach, spectral processing was investigated in R7/R8 inner photoreceptor terminals in the medulla. This study demonstrates that color opponency, a hallmark of spectral processing, is observable at the level of the first visual synapse in presynaptic terminals of the inner photoreceptors R7/R8. Two concurrent neuronal circuit mechanisms that involve distinct histamine receptors implement the comparison of R7 and R8 photoreceptor signals in the p and y pathways (Schnaitmann. 2018).

Recordings in norpA mutant flies combined with norpA rescue in pairwise combinations of photoreceptor types revealed mutual inhibitory interactions between R7 and R8 of the same type of ommatidia. These interactions provide the cellular basis for UVshort/blue and UVlong/green color-opponent responses in p and y photoreceptor terminals, respectively. Similar opponencies were previously posited to underlie color discrimination in dipteran flies). Thus, it is proposed that the photoreceptor terminals of the four types of inner photoreceptors mark the onset of two parallel color-opponent pathways that mediate color vision in Drosophila. The recently identified Tm5a/b/c, Tm9, Tm20, and TmY cells that are postsynaptic to R7/R8 likely represent further elements of these pathways). In particular, it has been suggested that Tm5a/b/c and Tm20 cells are elements of redundant color vision pathways. Blocking of all of these cell types, but not of single types or combinations, is required for complete loss of color discrimination. The finding of presynaptic color opponency in R7/R8 photoreceptors suggests that all of these neurons receive color-opponent input and do not generate color opponency de novo. These neurons likely participate in higher color processing, such as the spatial integration of spectral inputs, which is suggested for Tm5b/c cells based on their multi-columnar arborizations. Furthermore, synaptic connections from the outer photoreceptors R1-R6 to L3 lamina neurons and from L3 to some of the candidate neurons might explain the contribution of R1-R6 to color vision. At the level of R7/R8 terminals, no evidence was found for a role of R1-R6 photoreceptors in color-opponent processing (Schnaitmann. 2018).

The genetic, anatomical, and physiological experiments identify the two Drosophila histamine receptors HisCl1 and Ort) as key elements of direct and indirect inhibitory circuit mechanisms, respectively. In prior studies, chemical synaptic contacts of unknown function have been observed between R7/R8 photoreceptors by serial EM. The results on HisCl1 expression, and physiological recording suggest that these synapses are inhibitory and mediate color-opponent processing. Therefore, HisCl1 is of varying importance in R7 and R8 photoreceptors: in R7 terminals, hisCl1 expression is not required but is sufficient to generate color opponency; in R8 terminals, hisCl1 expression is required for color opponency, and it is only sufficient when ort expression in the visual circuits is intact. Interestingly, the varying importance of hisCl1-mediated direct inhibition between R7/R8 correlates well with the reported number of synapses between R7 and R8 photoreceptors. Serial EM reconstruction of one medulla column revealed 5 and 11 synapses from R7 onto R8 and vice versa, respectively (Schnaitmann. 2018).

Although ort is not expressed in photoreceptors, it nevertheless mediates color-opponent processing in concert with hisCl1. Physiological recordings reveal that ort is of varying importance for color opponency in R7 and R8, comparable with the results for hisCl1. Intact ort expression in the visual circuits is not required but sufficient for color-opponent processing in R7 terminals. In contrast, ort expression in the visual circuits is required for color opponency in R8 terminals, and it is only sufficient together with hisCl1 expression in R8 photoreceptor. The medulla neurons mediating ort-dependent feedback inhibition to R7/R8 terminals remain unknown. Among the many ort-expressing neurons in the medulla, several neuron types have been identified; however, none of these cell types establish feedback synapses onto R7/R8 photoreceptors. Work on polarization vision in flies has demonstrated that inner photoreceptors in the dorsal rim area similarly display antagonistic responses when the orientation of the e-vector is altered. If these inhibitory interactions are mediated by circuit mechanisms similar to the ones described in this study remains to be investigated. Along with the current results, the prior findings suggest that presynaptic calcium in Drosophila photoreceptor terminals is altered by local circuit interactions that enhance contrast (Schnaitmann. 2018).

Because the distal segments of the inner photoreceptors of large flies and of most other insects exhibit exclusively depolarizing non-opponent voltage responses, the observed color opponency in Drosophila inner photoreceptor terminals is unexpected, and it is proposed that spectrally antagonistic processing emerges only locally in photoreceptor terminals. Assuming that this local inhibitory signal does not propagate backward to the distal photoreceptor segments that have been recorded intracellularly, local processing would explain and reconcile the reported differences between insect species (Schnaitmann. 2018).

Recordings from R7/R8 terminals are consistent with mutually antagonistic processing of photoreceptors in the vertebrate retina. However, substantial differences in the molecular, synaptic, and network implementation demonstrate surprising variability in the underlying circuits (Schnaitmann. 2018).

Vertebrate photoreceptors employ cyclic guanosine monophosphate (cGMP) signaling, hyperpolarize, and reduce the release of glutamate in response to light. As a consequence, horizontal cells and OFF-bipolar cells expressing sign-conserving α-amino-3-hydroxy-5-methyl-4-isoxazol-propionacid (AMPA)/kainate-receptors receive less excitation, whereas ON-bipolar cells expressing sign-inverting metabotropic glutamate receptor 6 (mGluR6) receive less inhibition. In contrast, Drosophila photoreceptors employ phosphoinositide signaling and depolarize in response to light. This increases presynaptic calcium influx and the release of histamine, which binds to inhibitory Ort and HisCl1 receptors on postsynaptic cells. The results show that, in addition to interactions with Ort-expressing second-order neurons of the medulla, direct HisCl1-mediated interactions between inner photoreceptors play an important role in Drosophila color vision. Direct chemical synaptic interactions between different types of cones do not exist in the vertebrate retina. Vertebrate horizontal cells are the key players in early color-opponent processing: they receive input from different types of cones, synapse onto bipolar cells, and feed back onto cones with a sign-inverting synapse. Light-induced disinhibition of cone terminals is the fundamental mechanism underlying opponent L/M, and S/(L+M) interactions in cone terminals. If neurons analogous to horizontal cells exist in the Drosophila visual system has to be revealed. If so, they should participate in the Ort-dependent opponency mechanism revealed in this study. Based on the data and similar findings regarding the vertebrate retina, it is proposed that presynaptic color opponency in photoreceptor terminals is an important processing principle of color vision that is shared across taxa and that evolved in different taxa independently (Schnaitmann. 2018).

Inducible and titratable silencing of Caenorhabditis elegans neurons in vivo with histamine-gated chloride channels

Recent progress in neuroscience has been facilitated by tools for neuronal activation and inactivation that are orthogonal to endogenous signaling systems. This study describes a chemical-genetic approach for inducible silencing of Caenorhabditis elegans neurons in intact animals, using the histamine-gated chloride channel HisCl1 from Drosophila and exogenous histamine. Administering histamine to freely moving C. elegans that express HisCl1 transgenes in neurons leads to rapid and potent inhibition of neural activity within minutes, as assessed by behavior, functional calcium imaging, and electrophysiology of neurons expressing HisCl1. C. elegans does not use histamine as an endogenous neurotransmitter, and exogenous histamine has little apparent effect on wild-type C. elegans behavior. HisCl1-histamine silencing of sensory neurons, interneurons, and motor neurons leads to behavioral effects matching their known functions. In addition, the HisCl1-histamine system can be used to titrate the level of neural activity, revealing quantitative relationships between neural activity and behavioral output. These methods have been to dissect escape circuits, define interneurons that regulate locomotion speed (AVA, AIB) and escape-related omega turns (AIB), and demonstrate graded control of reversal length by AVA interneurons and DA/VA motor neurons. The histamine-HisCl1 system is effective, robust, compatible with standard behavioral assays, and easily combined with optogenetic tools, properties that should make it a useful addition to C. elegans neurotechnology (Pokala, 2014).

Histamine-HisCl1 receptor axis regulates wake-promoting signals in Drosophila melanogaster

Histamine and its two receptors, Histamine-gated chloride channel subunit 1 (HisCl1) and Ora transientless (Ort), are known to control photoreception and temperature sensing in Drosophila. However, histamine signaling in the context of neural circuitry for sleep-wake behaviors has not yet been examined in detail. Mutant flies were obtained with compromised or enhanced histamine signaling, and their baseline sleep was tested. Hypomorphic mutations in histidine decarboxylase (HDC), an enzyme catalyzing the conversion from histidine to histamine, caused an increase in sleep duration. Interestingly, hisCl1 mutants but not ort mutants showed long-sleep phenotypes similar to those in hdc mutants. Increased sleep duration in hisCl1 mutants was rescued by overexpressing hisCl1 in circadian pacemaker neurons expressing a neuropeptide pigment dispersing factor (PDF). Consistently, RNA interference (RNAi)-mediated depletion of hisCl1 in PDF neurons was sufficient to mimic hisCl1 mutant phenotypes, suggesting that PDF neurons are crucial for sleep regulation by the histamine-HisCl1 signaling. Finally, either hisCl1 mutation or genetic ablation of PDF neurons dampened wake-promoting effects of elevated histamine signaling via direct histamine administration. Taken together, these data clearly demonstrate that the histamine-HisCl1 receptor axis can activate and maintain the wake state in Drosophila and that wake-activating signals may travel via the PDF neurons (Oh, 2013).

Using genetic and pharmacological methods to manipulate histamine signaling, this study shows that the HisCl1 receptor and its downstream signaling cascade regulate wake-evoking behavior in Drosophila, while Ort receptor does not show any sleep/wake regulatory function. Histamine promotes activity via the HisCl1 receptor. Reduced histamine in HDC mutants or loss of the HisCl1 receptor both show reduced activity and enhanced sleep. Additionally, the relevant signaling pathway downstream of the HisCl1 receptor may function in the PDF neurons. Finally, it was demonstrated that the histamine-HisCl1 receptor axis can activate and maintain wakefulness in PDF neurons (Oh, 2013).

These data show the complete functional segregation of the two histamine receptors for the first time. Ort receptor is expressed in large monopolar cells (LMC), postsynaptic to photoreceptors in the lamina and is a major target of photoreceptor synaptic transmission in Drosophila. In contrast to Ort, HisCl1 receptor is not expressed in postsynaptic neurons of photoreceptors. It is expressed in lamina glia and shapes the LMC postsynaptic response of Ort signaling. Both Ort and HisCl1 receptor are involved in temperature-preference behaviors, but the major independent function of HisCl1 receptor remains elusive. This study showed that sleep regulation is a novel and independent function of HisCl1 receptor. Additionally, this finding is an important clue in understanding the functional evolution of the two histamine receptors in Drosophila (Oh, 2013).

It is proposed that wake-activation by histamine signaling in Drosophila is similar to that found in mammals. hdc mutant flies have increased sleep durations compared to controls and a previous study showed that HDC-knockout mice have increased paradoxical sleep compared to controls. This suggests that the HDC enzyme has a common wake-promoting function in mammals and flies. However, the structures of histamine receptors differ between flies and mammals; the histamine receptors of Drosophila are histamine-gated chloride channels, whereas the mammalian histamine receptors belong to the rhodopsin-like G-protein-coupled receptor family. Currently, researchers are working to identify a metabotropic histamine receptor in Drosophila. Despite the structural differences of the mammalian and Drosophila receptors, they share a wake-activating function. This functional homology may be the result of evolution and provides a hint to find out the metabotropic histamine receptors in Drosophila (Oh, 2013).

Surprisingly, functional conservations between flies and mammals are also found among the histamine receptor subtypes. The HisCl1 receptor has a wake-activating role, whereas the Ort receptor does not. This result parallels differences in the wake-activation roles of the H₁ and H₂ receptors in mammals: the H₁ receptor can activate wakefulness, but the H₂ receptor cannot. Thus, the data provide a more detailed understanding of the potential functional relationship between the HisCl1 and H₁ receptors. A functional connection between the Ort receptor and the H₂ receptor is also possible, since the two have little effect on sleep/wake regulation in their corresponding model systems. No auto-receptor of histamine has yet been found in Drosophila, suggesting that there may not be a Drosophila homolog for the mammalian H₃ receptor. Further research should shed greater light on the evolutionary relationship between the histamine receptors of flies and mammals (Oh, 2013).

Histamine signaling modulates the maintenance of wakefulness and controls light sensing, and it is speculated that a number of interactions are possible between these two different pathways. Previous studies on light-perception mechanisms showed that histamine mutants exhibit light-sensing defects. However, this study found that the sleep duration was increased in histamine signaling mutants compared to wild-type flies in constant darkness. Thus, the perception of light in the context of evoking wakefulness is independent of vision-related light perception in Drosophila. Further research will be required to definitively establish the relationship between light perception and sleep regulation (Oh, 2013).

Previous studies revealed that the PDF neurons promote wakefulness in Drosophila. The current findings show that histamine signaling acts as a wake-promoting pathway in PDF neurons. The HisCl1 receptor is a chloride channel, which would be expected to inhibit the function of the neurons. However, since previous studies showed that chloride channels can activate the function of the neurons, hence the HisCl1 receptor might be an activator of the PDF neurons. The downstream signaling of histamine-HisCl1 receptor in PDF neurons should be further studied using genetic manipulation and electro-physiological methods (Oh, 2013).

Orexin is a neuropeptide that acts as an important wake-activating neurotransmitter in mammals, as shown by the demonstration that defects in orexin synthesis can cause narcoleptic symptoms in human and animals. Orexin neurons activate wakefulness in the lateral hypothalamic area and the feedback loop between orexin neurons and monoaminergic neurons such as histaminergic and serotonergic neurons (tuberomammillary nucleus, TMN, and dorsal raphe nucleus, DR) controls wakefulness in the hypothalamus and the brain stem. Histamine receptors are essential for wake-activation by orexin treatment, indicating that orexin and histamine signaling constitute an interactive wake-activating system in mammals. However, orexin has not been found in Drosophila. A previous study suggested that the PDF neuropeptide functions similar to those of orexin in Drosophila , potentially explaining many aspects of the wake-activation cascade in Drosophila. Histamine and orexin have similar wake-activating function, but mammalian histamine mutants do not show narcoleptic symptoms. This study has shown that histamine and one of its receptors, HisCl1, constitute an important wake-evoking axis in Drosophila. Moreover, it was demonstrated that histamine-signaling mutants cannot maintain wakefulness during the daytime, which is similar to the phenotype of orexin mutants in mammals. Hence, it is proposed that, in Drosophila, histamine may have a function similar to that of the mammalian orexin. Further research is required to establish the functional relationship between wake activation of histamine signaling in Drosophila and wake-promoting function of orexin and histaminergic system in mammals (Oh, 2013).

Distinct roles for two histamine receptors (hclA and hclB) at the Drosophila photoreceptor synapse

Histamine (HA) is the photoreceptor neurotransmitter in arthropods, directly gating chloride channels on large monopolar cells (LMCs), postsynaptic to photoreceptors in the lamina. Two histamine-gated channel genes that could contribute to this channel in Drosophila are hclA (also known as ort) and hclB (also known as hisCl1), both encoding novel members of the Cys-loop receptor superfamily. Drosophila S2 cells transfected with these genes expressed both homomeric and heteromeric histamine-gated chloride channels. The electrophysiological properties of these channels were compared with those from isolated Drosophila LMCs. HCLA homomers had nearly identical HA sensitivity to the native receptors (EC(50) = 25 microM). Single-channel analysis revealed further close similarity in terms of single-channel kinetics and subconductance states (approximately 25, 40, and 60 pS, the latter strongly voltage dependent). In contrast, HCLB homomers and heteromeric receptors were more sensitive to HA (EC(50) = 14 and 1.2 microM, respectively), with much smaller single-channel conductances (approximately 4 pS). Null mutations of hclA (ort^US6096) abolished the synaptic transients in the electroretinograms (ERGs). Surprisingly, the ERG "on" transients in hclB mutants transients were approximately twofold enhanced, whereas intracellular recordings from their LMCs revealed altered responses with slower kinetics. However, HCLB expression within the lamina, assessed by both a GFP (green fluorescent protein) reporter gene strategy and mRNA tagging, was exclusively localized to the glia cells, whereas HCLA expression was confirmed in the LMCs. These results suggest that the native receptor at the LMC synapse is an HCLA homomer, whereas HCLB signaling via the lamina glia plays a previously unrecognized role in shaping the LMC postsynaptic response (Pantazis, 2008).

Histamine and its receptors modulate temperature-preference behaviors in Drosophila

Temperature profoundly influences various life phenomena, and most animals have developed mechanisms to respond properly to environmental temperature fluctuations. To identify genes involved in sensing ambient temperature and in responding to its change, >27,000 independent P-element insertion mutants of Drosophila were screened. As a result, it was found that defects in the genes encoding for proteins involved in histamine signaling [histidine decarboxylase (hdc), histamine-gated chloride channel subunit 1 (hisCl1), ora transientless (ort)] cause abnormal temperature preferences. The abnormal preferences shown in these mutants were restored by genetic and pharmacological rescue and could be reproduced in wild type using the histamine receptor inhibitors cimetidine and hydroxyzine. Spatial expression of these genes was observed in various brain regions including pars intercerebralis, fan-shaped body, and circadian clock neurons but not in dTRPA1-expressing neurons, an essential element for thermotaxis. The histaminergic mutants showed reduced tolerance for high temperature and enhanced tolerance for cold temperature. Together, these results suggest that histamine signaling may have important roles in modulating temperature preference and in controlling tolerance of low and high temperature (Hong, 2006).

Identification of two novel Drosophila melanogaster histamine-gated chloride channel subunits expressed in the eye

Histamine has been shown to play a role in arthropod vision; it is the major neurotransmitter of arthropod photoreceptors. Histamine-gated chloride channels have been identified in insect optic lobes. This study reports the first isolation of cDNA clones encoding histamine-gated chloride channel subunits from the fruit fly Drosophila melanogaster. The encoded proteins, HisCl1 and HisCl2, share 60% amino acid identity with each other. The closest structural homologue is the human glycine alpha3 receptor, which shares 45% and 43% amino acid identity respectively. Northern hybridization analysis suggested that hisCl1 and hisCl2 mRNAs are predominantly expressed in the insect eye. Oocytes injected with in vitro transcribed RNA, encoding either HisCl1 or HisCl2, produced substantial chloride currents in response to histamine but not in response to GABA, glycine, and glutamate. The histamine sensitivity was similar to that observed in insect laminar neurons. Histamine-activated currents were not blocked by picrotoxinin, fipronil, strychnine, or the H2 antagonist cimetidine. Co-injection of both hisCl1 and hisCl2 RNAs resulted in expression of a histamine-gated chloride channel with increased sensitivity to histamine, demonstrating coassembly of the subunits. The insecticide ivermectin reversibly activated homomeric HisCl1 channels and, more potently, HisCl1 and HisCl2 heteromeric channels (Zheng, 2002).

Search PubMed for articles about Drosophila HisCl1

Alejevski, F., Saint-Charles, A., Michard-Vanhee, C., Martin, B., Galant, S., Vasiliauskas, D. and Rouyer, F. (2019). The HisCl1 histamine receptor acts in photoreceptors to synchronize Drosophila behavioral rhythms with light-dark cycles. Nat Commun 10(1): 252. PubMed ID: 30651542

Hong, S. T., Bang, S., Paik, D., Kang, J., Hwang, S., Jeon, K., Chun, B., Hyun, S., Lee, Y., Kim, J. (2006). Histamine and its receptors modulate temperature-preference behaviors in Drosophila. J Neurosci, 26(27):7245-7256 PubMed ID: 23844178

Pantazis, A., Segaran, A., Liu, C. H., Nikolaev, A., Rister, J., Thum, A. S., Roeder, T., Semenov, E., Juusola, M., Hardie, R. C. (2008). Distinct roles for two histamine receptors (hclA and hclB) at the Drosophila photoreceptor synapse. J Neurosci, 28(29):7250-7259 PubMed ID: 24550306

Schnaitmann, C., Haikala, V., Abraham, E., Oberhauser, V., Thestrup, T., Griesbeck, O., Reiff, D. F. (2018). Color Processing in the Early Visual System of Drosophila. Cell, 172(1-2):318-330 e318 PubMed ID: 29328919

Zheng, Y., et al. (2002). Identification of two novel Drosophila melanogaster histamine-gated chloride channel subunits expressed in the eye. J. Biol. Chem. 277: 2000-2005. PubMed ID: 11714703

date revised: 17 April, 2024

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