A specific area of the compound eye in the cricket Gryllus bimaculatus sends photic information to the circadian pacemaker in the contralateral optic lobe

The circadian locomotor rhythm of the cricket Gryllus bimaculatus is primarily regulated by a pair of interacting optic lobe circadian pacemaker systems. The interaction involves phase-dependent modulation of the free-running period and phase-dependent suppression of activity. Since photic information has been shown to be involved in the interaction, we examined the regional difference in photoreception for the interaction within cricket compound eyes. The activity rhythm of animals receiving partial reduction of one compound eye combined with severance of the contralateral optic nerve split into entrained and free-running components under a 13-h light to 13-h dark cycle. All the animals operated on showed a phase-dependent suppression of activity, and most animals showed a phase-dependent modulation of the period of the free-running component. However, removal of the dorsocaudal area of the compound eye resulted in a severe reduction of the amplitude of the phase-dependent-period modulation. These results suggest that the dorsocaudal portion of the compound eye is a specific region receiving photic signals that are transmitted to the circadian pacemaker in the contralateral optic lobe and that the phase-dependent suppression of activity is caused by a mechanism separate from that for the period modulation.     

Isolation of an insect circadian clock

Summary Under constant conditions the compound eyes of the ground beetleAnthia sexguttata exhibit sensitivity changes in a very clear circadian rhythm. Usually the rhythms in both eyes in constant darkness are mutually coupled. After transection of the optic tract between the lobula and the supraesophageal ganglion the circadian rhythms of the two eyes continue without interruption, but coupling between them is abolished. Even if the entire supraesophageal ganglion is removed, leaving the optic ganglia intact, the circadian rhythms in the eyes continue without interruption independently. But the rhythm is abolished if the region of the lobula is damaged.The experiments show thatAnthia has circadian pacemakers in the left and right optic ganglia in or close to the lobula. These pacemakers can function independently from the rest of the brain and control circadian rhythms of physiological events.Supported by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 45 Vergleichende Neurobiologie des Verhaltens E1     

Circadian rhythms in the mating behavior of the cockroach, Leucophaea maderae

Mating behavior of small populations of virgin males and females of the cockroach Leucophaea maderae were continuously monitored via time-lapse video recording in controlled laboratory conditions. The time of onset of copulation was found to be rhythmic in a light cycle of 12 h light alternated with 12 h of darkness, with the peak of mating behavior occurring near the light to dark transition. This rhythm persisted in constant dim red illumination and constant temperature. In constant conditions, the period of the rhythm was slightly less than 24 h, with a peak of copulation during the late subjective day. These data demonstrated that mating behavior is gated by a circadian clock. When males and females were taken from light cycles that were 12 h out of phase, a bimodal rhythm was observed with one peak in the males' late subjective day and a second peak of equal amplitude in the late subjective day of females. The results indicated that circadian systems in both males and females contribute to the circadian rhythm in copulation. Bilateral section of the optic tracts (OTX) of both males and females abolished the rhythm, but the rhythm persisted when OTX females were paired with intact males or when OTX males were paired with intact females. Furthermore, when OTX males or OTX females were paired with intact animals that were 12 h out of phase, a bimodal rhythm was still observed. These results suggested that the circadian pacemaker in the optic lobes of both male and female cockroaches participates in the control of mating, but that a pacemaker outside the optic lobes is also likely involved. Finally, it was shown that the female's olfactory response (measured by electroantennogram) to components of the male sex pheromone exhibited a circadian rhythm, but the data suggested the peripheral olfactory rhythm is not likely to be involved in the rhythm of mating behavior.     

Localization of the circadian pacemakers of Hemideina thoracica (Orthoptera; Stenopelmatidae)

The location of the circadian pacemakers of the orthopteran Hemideina thoracica (White) has been investigated through observation of the effects of surgical removal of brain tissues (principally optic lobes and tracts) on free-running and entrained locomotor rhythms. Bilobectomy and severance of optic tracts invariably resulted in arrhythmicity, whereas rhythmicity was sustained following unilateral lobectomy, generally with increases in the free-running period (FRP) and decreases in both the active-phase lengths and activity-to-rest ratios of the rhythm. Bilobectomized subjects could be entrained by temperature cycles, but exhibited no transients or residual rhythmicity, indicating that temperature brought about a direct response or masking effect. These results support the hypothesis that the circadian locomotor pacemakers of Hemideina are located within each optic lobe, and that there are no extraoptic centers for the control of the timing of locomotor activity. Although confirmation of the pacemaker role of the optic lobes requires transplantation of the tissues, the conclusion may be drawn by inference from other studies (e.g., Leucophaea maderae--Page, 1983; Gryllus bimaculatus--Tomioka and Chiba, 1986). Light entrainment continued after surgical binding and blackening of the compound eyes and ocelli, supporting the view that direct illumination of neural tissue through the cuticle may be one possible pathway for light entrainment.     

Scorpion lateral eyes: Extremely sensitive receptors of Zeitgeber stimuli

Summary The circadian rhythm of sensitivity in the median eyes ofAndroctonus australis L. can be entrained by exposure of the lateral eyes to a 24-h light-dark rhythm. Presentation of the Zeitgeber to only the anteriormost one of the lateral eyes sufficed (Fig. 1). However, with illumination of an entire group of lateral eyes (Fig. 2), entrainment was obtained at extremely low light intensities — white light at luminance levels of 10^–4cd · m^–2 (=2.5 · 10^–4 lux, cf. Methods).The relatively less marked circadian rhythm of lateral-eye sensitivity is probably controlled via the optic nerve supplying these eyes (Fig. 4).Supported by the Deutsche Forschungsgemeinschaft (F1 77/5-6 and F1 77/7 Schwerpunktprogramm: Biologie der Zeitmessung)     

Odor discrimination using insect electroantennogram responses from an insect antennal array

Insects have a highly developed olfactory sensory system, mainly based in their antennae, for the detection and discrimination of volatile compounds in the environment. Electroantennogram (EAG) response profiles of five different insect species, Drosophila melanogaster, Heliothis virescens, Helicoverpa zea, Ostrinia nubilalis and Microplitis croceipes, showed different, species-specific EAG response spectra to 20 volatile compounds tested. The EAG response profiles were then reconstructed for each compound across the five insect species. Most of the compounds could be distinguished by comparing the response spectra. We then used a four-antenna array, called a Quadro-probe EAG, to see if we could discriminate among odorants based on the relative EAG amplitudes evoked when the probe was placed in plumes in a wind tunnel and in a field. Stable EAG responses could be simultaneously and independently recorded with four different insect antennae mounted on the Quadro-probe, and different volatile compounds could be distinguished in real time by comparing relative EAG responses with a combination of differently tuned insect antennae. Regardless of insect species or EAG amplitudes, antennae on the Quadro-probe maintained their responsiveness with higher than 1 peak/s of time resolution.     

Protein synthesis is a required process for the optic lobe circadian clock in the cricket Gryllus bimaculatus

The effects of a translation inhibitor, cycloheximide (CHX), on the circadian neuronal activity rhythm of the optic lamina-medulla compound eye complex cultured in vitro were investigated in the cricket Gryllus bimaculatus. When the complex was treated with 10(-5) M CHX for 6 h, the rhythm exhibited a marked phase shift. The magnitude and direction of the phase shift were dependent on the phase at which the complex was treated with CHX; phase delays occurred during the late subjective day to early subjective night, whereas phase advances occurred around the late subjective night. Continuous application of CHX abolished circadian rhythms of both the spontaneous neuronal activity and the visually evoked response. However, it abolished neither the spontaneous activity nor the visually evoked response. As washed with fresh medium after CHX treatment, the rhythm soon reappeared and the subsequent phase was clearly correlated to the termination time of the treatment. These results suggest that protein synthesis is also involved in the cricket optic lobe circadian clock, and that the clock-related protein synthesis may be active during the late subjective day to subjective night.     

Distribution of circadian photoreceptors in the compound eye of the cricket Gryllus bimaculatus

Adult male crickets (Gryllus bimaculatus) show a nocturnal circadian locomotor rhythm, which is driven by the pacemaker in the optic lamina-medulla complex and synchronizes to the light-dark (LD) cycle received by the compound eye. To see whether there was any specially differentiated circadian photoreceptor area in the eye, we examined the effect of a partial reduction of various areas of the compound eye, in addition to a removal of the contralateral optic lamina-medulla-compound eye complex, on entrainability of the locomotor rhythm. All operated animals showed a response to the LD cycle in their locomotor rhythm, no matter which area of the eye was left intact: They either stably entrained to an LD cycle or showed a sign of weak entrainment. The capacity for stable entrainment was still retained when only 262 ommatidia were left. Transient cycles needed for re-entrainment, following a 6-hr phase advance of the LD cycle, were measured in 20 reduced-eye animals showing clear stable entrainment. They were in inverse proportion to the number of ommatidia in the reduced eye: The fewer ommatidia there were, the more transient cycles were observed (r = -0.76, p less than 0.001). These results suggest that almost the whole area of the compound eye may contain circadian photoreceptors, and that the photic information from each ommatidium may additively affect the circadian clock to entrain via neural integration mechanisms.     

Neural pathways involved in mutual interactions between optic lobe circadian pacemakers in the cricketGryllus bimaculatus

The circadian locomotor rhythm of the cricketGryllus bimaculatus is primarily generated by a pair of optic lobe circadian pacemakers. The two pacemakers mutually interact to keep a stable temporal structure in the locomotor activity. The interaction has two principal effects on the activity rhythm, i.e., phase-dependent modulation of the freerunning period and phase-dependent suppression of activity driven by the partner pacemaker. Both effects were mediated by neural pathways, since they were immediately abolished after the optic stalk connecting the optic medulla to the lobula was unilaterally severed. The neural pathways were examined by recording locomotor activity, under a 13 h light to 13 h dark cycle, after the optic nerves were unilaterally severed and the contralateral optic stalk was partially destroyed near the lobula. When the dorsal half of the optic stalk was severed, locomotor rhythm mostly split into two components: one was readily entrained to the given light-dark cycle and the other freeran with a marked fluctuation in freerunning period, where the period of the freerunning component was lengthened or shortened when the onset of the entrained component occurred during its subjective night or day, respectively. The phase-dependent modulation of activity was also observed in both components. However, severance of the ventral half of the optic stalk resulted in appearance only of the freerunning component; neither the phase-dependent modulation of its freerunning period nor the change in activity level was observed. These results suggest that neurons driving the mutual interaction and the overt activity rhythm run in the ventral half of the proximal optic stalk that includes axons of large medulla neurons projecting to the cerebral lobe and the contralateral medulla.Abbreviations LD light dark cycle - freerunning period     

Circadian rhythms in the electroretinogram of the cockroach

Circadian regulation of the amplitude of the electroretinogram (ERG) of the cockroach Leucophaea maderae was investigated. Two components of the ERG exhibited circadian rhythms in amplitude. Interestingly, the peak amplitudes for the two rhythms were approximately 12 hr out of phase. The dominant corneal negative potential (the "sustained component") exhibited maximum amplitude during the subjective night. A second corneal negative potential (the "off-transient") was at a maximum during the subjective day. Intensity-response curves of the sustained component were measured at both the peak and trough of the rhythm. The results showed that the circadian rhythm in amplitude reflected a sensitivity change equivalent to 0.2-0.6 log unit of intensity. An effort was also made to identify the anatomical locus of the pacemaking oscillator for the ERG rhythm in a series of lesion experiments. Neural isolation of the optic lobe from the midbrain by bisection of the optic lobe proximal to the distal edge of the lobula had no effect on the circadian rhythm of ERG amplitude. Bisection of the optic lobe distal to the lobula abolished the ERG amplitude rhythm. These results suggest that the pacemaker is located in the optic lobe near the lobula; that its motion continues in the absence of neural connections with the rest of the nervous system; and that its regulation of ERG amplitude depends on neural pathways in the optic lobe.     

Drosophila olfactory response rhythms require clock genes but not pigment dispersing factor or lateral neurons

Odors elicit a number of behavioral responses, including attraction and repulsion in Drosophila. In this study, the authors used a T-maze apparatus to show that wild-type Drosophila melanogaster exhibit a robust circadian rhythm in the olfactory attractive and repulsive responses. These responses were lower during the day and began to rise at early night, peaking at about the middle of the night and then declining thereafter. They were also independent of locomotor activity. The olfactory response rhythms were lost in period or timeless mutant flies (per0, tim0), indicating that clock genes control circadian rhythms of olfactory behavior. The rhythms in olfactory response persisted in the absence of the pigment-dispersing factor neuropeptide or the central pacemaker lateral neurons known to drive circadian patterns of locomotion and eclosion. These results indicate that the circadian rhythms in olfactory behavior in Drosophila are driven by pacemakers that do not control the rest-activity cycle and are likely in the antennae.     

The circadian rhythm of the sex-pheromone-mediated behavioral response in the turnip moth, Agrotis segetum, is not controlled at the peripheral level

The pheromone-mediated upwind flight of male turnip moths was observed in a flight tunnel at different times of day under conditions of a light-dark (LD) cycle, constant darkness (DD), and a shifted photoperiod. Under both LD and DD conditions, a significantly larger number of males flew to the pheromone during both the scotophase and the subjective scotophase than during the photophase and the subjective photophase for 2 consecutive days. When 1-day-old moths were transferred to a shifted LD cycle with lights turned off 4 h earlier, male behavioral responses to the pheromone advanced in time accordingly by 4 h. This showed that male behavioral responses to the pheromone are under the control of an endogenous oscillator. To further examine the level at which the circadian rhythm of the male behavioral response is regulated, the authors tested the olfactory responses of male antennal receptors to pheromone stimuli by means of electroantennograms (EAG) at different times of day. No significant variation in the sensitivity of the male antennal response to the pheromone was observed in terms of time of day. The results suggest that circadian regulation of the rhythmic behavioral response to pheromones in the male Agrotis occurs at the central nervous system level.     

性信息素对斜纹夜蛾雄蛾嗅觉相关基因 abp,pbp 和 or 表达的影响

【目的】信息素是个体之间传递信息的重要分子,研究性信息素对斜纹夜蛾 Spodoptera litura 嗅觉相关基因表达的影响对于增加性信息素作用机理的认识及其应用有重要的意义。【方法】本研究通过实时定量PCR（qRT-PCR）技术探究在性信息素刺激处理条件下,斜纹夜蛾成虫嗅觉相关基因 abp, pbp 和 or 表达水平的变化;利用性信息素在田间诱捕斜纹夜蛾雄蛾,并通过自动计数器记录每小时诱虫量,从而间接显示其交配行为的节律性。【结果】斜纹夜蛾雄虫触角中嗅觉相关基因abp, pbp 和 or 的表达具有节律特性。经性信息素化合物(Z9, Z11-14:OAc+Z9, Z12-14:OAc)刺激处理后,abp, pbp 和 or 表达量也发生了显著的改变。通过记录田间性信息素诱捕器在一天中不同时间段内诱捕的雄蛾数量,发现诱捕到的斜纹夜蛾也具有节律特性。【结论】基因表达水平上的节律特性可能与雄虫交配活动的节律相关联,说明性信息素处理也在一定程度上改变了其节律及其对性信息素的神经反应。这一结果也首次从基因水平证明性信息素的刺激处理提高了周缘神经系统对性信息素反应的敏感性,有助于我们理解性信息素作用的分子机理,对迷向及性诱和测报应用具有指导意义。     

Improvement of signal-to-noise ratio in electroantennogram responses using multiple insect antennae

Using an array of insect antennae connected in series or in parallel, electroantennogram (EAG) responses and noise levels were investigated in an attempt to improve signal-to-noise (S/N) ratio and sensitivity. Both the EAG response amplitude and noise level increased when the antennae of male Helicoverpa zea (Lepidoptera: Noctuidae) were connected in series. Due to lower relative increase in noise level than EAG amplitude as the number of antennae increased, the S/N ratio was also significantly improved by the serial connection. As a result the sensitivity of EAG was improved by the serial connection, which showed ca. ten-fold improvement in the threshold detection levels compared with a single antenna when four antennae were connected in series. In contrast to the serial connection, there were no differences in EAG amplitudes and overall noise levels when different numbers of antennae were connected in parallel. When only large-amplitude noise was taken into account, however, the S/N ratio was somewhat improved by the parallel connection. The frequency of overall noise remained at the same level both in the serial and in the parallel connection. However, the frequency of the large-amplitude noise increased in serial connection but decreased in parallel connection. The present study clearly indicates that both the sensitivity and S/N ratio of the EAG biosensor could be significantly improved by using the multiple antennal connections.     

Morphological and functional heterogeneity in olfactory perception between antennae and maxillary palps in the pumpkin fruit fly,Bactrocera depressa

The morphology and ultrastructure of the olfactory sensilla on the antennae and maxillary palps were investigated through scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and their responses to five volatile compounds were measured using electroantenogram (EAG) and electropalpogram (EPG) techniques in the pumpkin fruit fly, Bactrocera depressa (Shiraki; Diptera: Tephritidae). Male and female B. depressa displayed distinct morphological types of olfactory sensilla in the antennae and maxillary palps, with predominant populations of trichoid, basiconic, and coeloconic sensilla. Basiconic sensilla, the most abundant type of olfactory sensilla in the antennae, could be further classified into two different types. In contrast, the maxillary palps exhibited predominant populations of a single type of curved basiconic sensilla. High‐resolution SEM observation revealed the presence of multiple nanoscale wall‐pores on the cuticular surface of trichoid and basiconic sensilla, indicating that their primary function is olfactory. In contrast, coeloconic sensilla displayed several longitudinal grooves around the sensillum peg. The TEM observation of individual antennal olfactory sensilla indicates that the basiconic sensilla are thin‐walled, while the trichoid sensilla are thick‐walled. The profile of EAG responses of male B. depressa was different from their EPG response profile, indicating that the olfactory function of maxillary palps is different from that of antennae in this species. The structural and functional variation in the olfactory sensilla between antennae and maxillary palps suggests that each plays an independent role in the perception of olfactory signals in B. depressa.     

Phototaxis by the Dinoflagellate Gymnodinium splendens Lebour*

A microscope-television system was used to monitor quantitatively the behavior of Gymnodinium splendens Lebour in response to light. The predominant behavioral sequence upon stimulation is (a) an initial 2–5 sec cessation of movement (stop-response) followed by (b) positive phototaxis. The action spectra for each response are identical, having maxima at 450 and 280 nm. Upon measuring the percent response to a range of stimulus intensities, it is apparent that a stop-response is not a behavioral prerequisite for phototaxis. An identical circadian rhythm in photoresponsiveness is observed for phototaxis and for the stop-response with greatest light sensitivity occurring during the first 4 hr of the entrained light period. The implication of phototactic sensitivity and the phototactic circadian rhythm in diurnal vertical migration is discussed.     

Effects of optic lobe ablation on circadian activity in the mosquito, Culex pipiens pallens

ABSTRACT. Surgical manipulation was used to show that in the mosquito, Culex pipiens pallens , the circadian pacemaker is probably not located in the optic lobes. Mosquitoes deprived of the greater part of their optic lobes still maintained a circadian activity rhythm: in DD with τ= c . 20–23 h, in LL with τ= c . 14–15 h. The operated mosquitoes showed diphasic activity which entrained to LD 16:8 h (at 200 lux), implying the existence of an extraocular photoreceptive pathway.     

An improved aphid electroantennogram

Excised antennae have been used to record aphid electroantennograms (EAGs) but these preparations have small, rapidly declining responses and a short usable life. An improved EAG technique is described and evaluated using alate virginoparae of the black bean aphid, Aphis fabae, and a series of plant volatiles. EAG recording with fine-tip glass electrodes inserted into the surface of the intact antenna of a whole insect preparation gave stable EAG peaks of a few mV and EAG peak amplitude decreased by only 22% over an 8 hour recording period. The EAG responses showed typical dose-dependent characteristics. There was variation in EAGs from different preparations, so normalisation of the EAG responses against a standard stimulus is still necessary. The thirty compounds tested elicited an EAG response profile largely similar to that reported previously from excised antennae. EAG responses recorded with the electrode at three different positions, between the 5th-6th (A), 4th-5th (B), and 3rd-4th (C) antennal segments, were smaller when recorded closer to the head. Position B produced larger EAG responses than those at C although there was no olfactory receptor between position B and C.     

Manipulating the light/dark cycle: effects on dopamine levels in optic lobes of the honey bee (<Emphasis Type="Italic">Apis mellifera</Emphasis>) brain

This study examines the relationship between cyclical variations in optic-lobe dopamine levels and the circadian behavioural rhythmicity exhibited by forager bees. Our results show that changing the light–dark regimen to which bees are exposed has a significant impact not only on forager behaviour, but also on the levels of dopamine that can be detected in the optic lobes of the brain. Consistent with earlier reports, we show that foraging behaviour exhibits properties characteristic of a circadian rhythm. Foraging activity is entrained by daily light cycles to periods close to 24 h, it changes predictably in response to phase shifts in light, and it is able to free-run under constant conditions. Dopamine levels in the optic lobes also undergo cyclical variations, and fluctuations in endogenous dopamine levels are influenced significantly by alterations to the light/dark cycle. However, the time course of these changes is markedly different from changes observed at a behavioural level. No direct correlation could be identified between levels of dopamine in the optic lobes and circadian rhythmic activity of the honey bee.