Circadian Rhythmicity in Excised Samanea Pulvini: II. Resetting the Clock by Phytochrome Conversion

Excised Samanea saman pulvini were incubated in H(2)O or 50 mm sucrose in darkness for 100 to 152 hours except for brief exposures to red or far red light, and angles of opening measured periodically. When pulvini are incubated in H(2)O, the rhythm damps in the open position after two to three cycles irrespective of the light treatments, but when sucrose is available, the now persistent oscillations show large red, far red-regulated effects on phase, amplitude, mesor slope, and entrainment. Single red light pulses rephase the rhythm, with a phase response curve that resembles that reported for other plants and animals; such rephasing is prevented by immediately subsequent far red light, indicating that phytochrome is the photoreceptor. Red light pulses repeated every 24 hours entrain the rhythm, and also prevent damping if presented at an appropriate part of the cycle.     

A circadian rhythm in oxygen uptake by samanea pulvini

The rate of O₂ uptake by excised Samanea pulvini oscillates with a circadian rhythm during 52 hours of darkness. Rates of respiration increase during pulvinar opening and decrease prior to and during closure, consistent with the concept that opening requires a greater expenditure of energy. Externally supplied sucrose, necessary for perpetuation of the leaflet movement rhythm, has a small promotive effect on the rate of respiration.     

Relationships between Motor Cell Ultrastructure and Leaf Movements in Samanea saman

We examined the fine structure of motor cells in the secondary pulvinus of Samanea saman (Jacq.) Merrill, to aid in analyzing the cellular basis for K⁺ and Cl^? driven, turgor regulated circadian leaflet movements. Pulvini that were (a) open (horizontal) in the light, (b) closed (vertical) in the dark, or (c) at an intermediate angle after 96-h incubation in H₂O in darkness, were cut into cross sections, sub-divided into quadrants, and prepared for electron microscopy by standard methods. The walls of many cells are ridged, appearing scalloped in cross section, the plasmalemma following the wall contours. Plasmodesmata localized in pit fields are most numerous in the inner cortex of the extensor (abaxial), and least numerous in the outer cortex of the flexor (adaxial) (3.9 and 0.7, respectively, per μm² wall area). Vacuole size, number of vacuoles per cell, and the condition of the electron dense precipitate within the vacuole also vary with cell location, multivacuolation being most pronounced in the outer cortex of the extensor. Chloroplasts are dimorphic: those in cells close to the vascular tissue are very large, circular in cross section, and contain huge starch deposits at all times, while those in the remainder of the cortex are smaller, usually oblong, and contain large starch deposits at the beginning of the dark period, but are devoid of starch at the beginning of the light period. However, both types of chloroplasts in excised pulvini incubated in H²O in darkness for 96 h still contain starch deposits, indicating that (1) light may promote starch degradation, or (2) starch degradation and resynthesis may be rhythmic.     

Effect of light during lactation on the phasic and tonic responses of the rat pacemaker

The circadian system in mammals generates endogenous circadian rhythms and entrains them to external cycles. Here, we examine whether the lighting conditions under which rats are reared affect the properties of the circadian pacemaker. We maintained three groups of rats under constant darkness (DD-rats), constant bright light (LL-rats) or light-dark cycles of 24 hours (LD-rats) during lactation. We then studied motor activity rhythm under constant light of four intensities, and under seven light-dark cycles with periods ranging between 22 and 27 hours. Results show that neither the tau nor the phase angle to the external cycle differed between groups. Differences were found in the amplitude of the circadian rhythm and in the number of rats that became arrhythmic under LL. We conclude that the light received during lactation affects the strength of the circadian pacemaker and its sensitivity to light.     

Latitudinal variation in eclosion rhythm among strains of Drosophila ananassae.

Eclosion rhythm parameters of D. ananassae strains originating between 8 degrees-34 degrees N were highly variable and latitude dependent. In the field under naturally fluctuating light intensity, temperature and R.H., the amplitude of the rhythm was high and the eclosion gate was narrow; however, under the naturally fluctuating light intensity but at constant temperature and R.H., the amplitude of the rhythm was lowered and the width of eclosion gate was widened. The eclosion rhythm entrained to light-dark (LD) cycles ranging from LD 6:18 to LD 18:6, the width of the eclosion gate was decreased and increased in the short and long photoperiods respectively. Among the strains, both the phase angle difference (psi, the time from lights-off in a 24 hr LD cycle to the eclosion median) and the period of free-running rhythm (tau) in constant darkness varied by about 3 hr and the amplitude of the rhythmicity (Amp) by about 10%. Lower latitude was correlated with late psi (r = -0.69), long tau (r = -0.88) and high Amp value (r = -0.95).     

Light pulses entrain the circadian activity rhythm of a diurnal rodent (Ammospermophilus leucurus)

Circadian rhythms of wheel-running activity of the antelope ground squirrel (Ammospermophilus leucurus) were entrained by light-dark cycles (LD: 100 1x vs total darkness) with periods (T) between ca 23.75 and 24.75 hr. Two 1-hr light pulses per cycle ('skeleton photoperiods') with T = 24.25 hr as well as one 1-hr light pulse per cycle with Ts of 23.75 and 24.25 hr were effective in entraining the circadian activity rhythms in at least 50% of the antelope ground squirrels. Phase and period responses to single 1-hr light pulses were measured which depend on the initial phase and period of the rhythm. It is concluded that discrete (phasic) light input contributes to the mechanism of entrainment to LD cycles in diurnal rodents.     

Effects of constant darkness and constant light on circadian organization and reproductive responses in the ram

The relationship between circadian rhythms in the blood plasma concentrations of melatonin and rhythms in locomotor activity was studied in adult male sheep (Soay rams) exposed to 16-week periods of short days (8 hr of light and 16 hr of darkness; LD 8:16) or long days (LD 16:8) followed by 16-week periods of constant darkness (dim red light; DD) or constant light (LL). Under both LD 8:16 and LD 16:8, there was a clearly defined 24-hr rhythm in plasma concentrations of melatonin, with high levels throughout the dark phase. Periodogram analysis revealed a 24-hr rhythm in locomotor activity under LD 8:16 and LD 16:8. The main bouts of activity occurred during the light phase. A change from LD 8:16 to LD 16:8 resulted in a decrease in the duration of elevated melatonin secretion (melatonin peak) and an increase in the duration of activity corresponding to the changes in the ratio of light to darkness. In all rams, a significant circadian rhythm of activity persisted over the first 2 weeks following transfer from an entraining photoperiod to DD, with a mean period of 23.77 hr. However, the activity rhythms subsequently became disorganized, as did the 24-hr melatonin rhythms. The introduction of a 1-hr light pulse every 24 hr (LD 1:23) for 2 weeks after 8 weeks under DD reinduced a rhythm in both melatonin secretion and activity: the end of the 1-hr light period acted as the dusk signal, producing a normal temporal association of the two rhythms. Under LL, the 24-hr melatonin rhythms were disrupted, though several rams still showed periods of elevated melatonin secretion. Significant activity rhythms were either absent or a weak component occurred with a period of 24 hr. The introduction of a 1-hr dark period every 24 hr for 2 weeks after 8 weeks under LL (LD 23:1) failed to induce or entrain rhythms in either of the parameters. The occurrence of 24-hr activity rhythm in some rams under LL may indicate nonphotoperiodic entrainment signals in our experimental facility. Reproductive responses to the changes in photoperiod were also monitored. After pretreatment with LD 8:16, the rams were sexually active; exposure to LD 16:8, DD, or LL resulted in a decline in all measures of reproductive function. The decline was slower under DD than LD 16:8 or LL.(ABSTRACT TRUNCATED AT 400 WORDS)     

CIRCADIAN GROWTH RHYTHM IN JUVENILE SPOROPHYTES OF LAMINARIALES (PHAEOPHYTA)1

A circadian rhythm in growth was detected by computer-aided image analysis in 3–4-cm-long, juvenile sporophytes of the kelp species Pterygophora California Rupr. and in seven Laminaria spp. In P. californica, the free-running rhythm occurred in continuous white fluorescent light, had a period of 26 h at 10°or 15°C, and persisted for at least 2 weeks in white or blue light. The rhythm became insignificant in continuous green or red light after 3 cycles. Synchronization by white light-dark regimes, e.g. by 16 h light per day, resulted in an entrained period of 24 h and in a shift of the circadian growth minimum into the middle of the light phase. A morning growth peak represented the decreasing portion of the circadian growth curve, and an evening peak the increasing portion. The circadian growth peak was not visible during the dark phase, because growth rate decreased immediately after the onset of darkness. At night, some growth still occurred at 16 or 12 h light per day, whereas growth stopped completely at 8 h light per day, as in continuous darkness. During 11 days of darkness, the thallus area became reduced by 3.5%, but growth rate recovered in subsequent light–dark cycles, and the circadian growth rhythm reappeared in subsequent continuous light.     

Circadian phototransduction: phase resetting and frequency of the circadian clock of Gonyaulax cells in red light

Constant red light (RR) influences the Gonyaulax clock in several ways: (1) Phase resetting by white or blue light pulses is stronger under background RR than in constant white light (WW); (2) frequency of the rhythm is less in RR than in WW; and (3) the amplitude of the spontaneous flashing rhythm is greater in RR than in WW. The phase response curve (PRC) to 4-hr white or blue light pulses is of high amplitude (Type 0) for cells in RR, but is of lower amplitude (Type 1) for cells in WW. In all cases, the PRC is highly asymmetrical: The magnitude of advance phase resetting is far higher than that of delay resetting. Consistent with this PRC, Gonyaulax cells in RR (free-running period greater than 24 hr) will entrain to T cycles of between 21 and 26.5 hr. The bioluminescence rhythms exhibit "masking" by blue light pulses while entrained to these T cycles. The fluence response of phase resetting to light-pulse intensity is not linear or logarithmic--rather, it is discontinuous. This feature is consistent with a limit cycle interpretation of Type 0 resetting of circadian clocks. Light pulses that cause large phase shifts also shorten the subsequent free-running period. The phase angle difference between the clock and the previous LD cycle is within 2 hr of the same phase between 16 degrees C and 25 degrees C, as determined from the light PRCs at various temperatures. Several drugs that inhibit mitochondria and/or electron transport will partially inhibit the phase shift by light.     

Unterschiedliche Lichtintensitätsabhängigkeit der Spontanperioden als Ursache interner Desynchronisation circadianer Rhythmen bei Phaseolus coccineus

Summary The length of the free-running periods of circadian leaf movements in the primary and secondary pulvini of Phaseolus coccineus was measured in constant darkness (DD) and continuous light (LL) of different intensities. The periods for the two pulvini do not differ from each other in DD and LL 30000 lx. However, they differ in LL 60 lx, 600 lx and 6000 lx, indicating a state of internal desynchronization. These results show a difference in the light intensity dependence of the 2 oscillations and a lack of mutual synchronization between them.     

Effects of Light on the Membrane Potential of Protoplasts from Samanea saman Pulvini : Involvement of K Channels and the H -ATPase

Rhythmic light-sensitive movements of the leaflets of Samanea saman depend upon ion fluxes across the plasma membrane of extensor and flexor cells in opposing regions of the leaf-movement organ (pulvinus). We have isolated protoplasts from the extensor and flexor regions of S. saman pulvini and have examined the effects of brief 30-second exposures to white, blue, or red light on the relative membrane potential using the fluorescent dye, 3,3′-dipropylthiadicarbocyanine iodide. White and blue light induced transient membrane hyperpolarization of both extensor and flexor protoplasts; red light had no effect. Following white or blue light-induced hyperpolarization, the addition of 200 millimolar K⁺ resulted in a rapid depolarization of extensor, but not of flexor protoplasts. In contrast, addition of K⁺ following red light or in darkness resulted in a rapid depolarization of flexor, but not of extensor protoplasts. In both flexor and extensor protoplasts, depolarization was completely inhibited by tetraethylammonium, implicating channel-mediated movement of K⁺ ions. These results suggest that K⁺ channels are closed in extensor plasma membranes and open in flexor plasma membranes in darkness and that white and blue light, but not red light, close the channels in flexor plasma membranes and open them in extensor plasma membranes. Vanadate treatment inhibited hyperpolarization in response to blue or white light, but did not affect K⁺ -induced depolarization. This suggests that white or blue light-induced hyperpolarization results from activation of the H⁺ -ATPase, but this hyperpolarization is not the sole factor controlling the opening of K⁺ channels.     

Evidence of phytochrome involvement in the entrainment of the circadian rhythm of carbon dioxide metabolism in Bryophyllum

The rhythm of carbon dioxide output in Bryophyllum leaves was entrained on exposure to 0.25 h of white light every 24 h. Entrainment also occurred on similar exposure to monochromatic radiation in spectral bands centred at 660 nm and, to a lesser extent, at 730 nm, but a band centred at 450 nm was without effect. A skeleton irradiation programme comprising two 0.25-h exposures to white light per 24 h also entrained the rhythm when the intervening dark periods were either 7.5 h and 16 h, or 10.5 h and 13 h. The rhythm disappeared when the two exposures were separated by 11.5-h and 12-h dark periods. Regular 0.25-h exposures to red light separated by 11.75-h periods of darkness also resulted in loss of the rhythm. Red/far-red reversibility was observed in irradiation schedules having either one or two exposures to red light daily. In the latter case, far-red reversal of the effects of one of the exposures to red light resulted in entrainment of the rhythm by the other, instead of abolition of the rhythm. The occurrence of distinct red/far-red reversibility suggests strongly that phytochrome is the pigment involved in entrainment of this rhythm by cycles of light and darkness.Abbreviation LD light-dark rhythm     

Different circadian rhythms regulate photoperiodic flowering response and leaf movement in Pharbitis nil (L.) Choisy

The phase shifting effect of red light on both the leaf movement rhythm, and on the rhythm of responsiveness of photoperiodic flower induction towards short light breaks (10 min red light), has been studied in Pharbitis nil, strain Violet, and comparisons between the two rhythms have been made. The phase angle differences between the rhythms after a phase shift with 2 or 6 h of red light given at different times during a long dark period were not constant. The results indicate the involvement of two different clocks controlling leaf movement and photoperiodic flower induction.Abbreviations DD continuous darkness - l:D x:y light/dark cycles with x hours of light and y hours of darkness - PPR rhythm of photoperiodic responsiveness towards light break     

Potassium Flux and Leaf Movement in Samanea saman : I. Rhythmic Movement

Samanea leaflets usually open in white light and fold together when darkened, but also open and dose with a circadian rhythm during prolonged darkness. Leaflet movement results from differential changes in the turgor and shape of motor cells on opposite sides of the pulvinus; extensor cells expand during opening and shrink during closure, while flexor cells shrink during opening and expand during closure but change shape more than size. Potassium in both open and closed pulvini is about 0.4 N. Flame photometric and electron microprobe analyses reveal that rhythmic and light-regulated postassium flux is the basis for pulvinar turgor movements. Rhythmic potassium flux during darkness in motor cells in the extensor region involves alternating predominance of inwardly directed ion pumps and leakage outward through diffusion channels, each lasting ca 12 h. White light affects the system by activating outwardly directed K⁺ pumps in motor cells in the flexor region.     

Circadian control of spontaneous flight activity in the blowfly, Lucilia cuprina

ABSTRACT. Under laboratory light: dark cycles, the flight activity of adult Lucilia cuprina (Wied.) was low during darkness and uniformity high during light. This pattern persisted as a rhythm both in constant darkness and in constant light of intensity up to 1lx, with a period of approximately 22 h in each. Light pulses of 15 min at l00lx applied to the free-running rhythm in constant darkness generated phase shifts of up to 60°, 12-h light pulses of the same intensity generated maximal (180°) phase shifts. The phase response curves had shapes similar to those of a number of other insect rhythms. When exposed to light periods (70 lx) of greater than 12 h followed by constant darkness, the rhythm reinitiated at the light-dark transition from a constant phase equivalent to that at the time of the light-dark transition in the LD 12:12 cycle.     

Daily light regulates seasonal responses in the migratory male redheaded bunting (Emberiza bruniceps)

This study analyzed the role of day length in regulation of seasonal body fattening and testicular growth in a latitudinal Palaearctic-Indian migrant, the redheaded bunting (Emberiza bruniceps). When exposed to increasing photoperiods (hours of light: hours of darkness; 11.5L:12.5D, 12L:12D, 12.5L:11.5D, 13L:11D, 14L:10D, and 18L:6D) for 9-12 weeks, buntings responded in a photoperiod-dependent manner and underwent growth and regression cycle under photoperiods of > or =12 hr per day. Also, the response to a long photoperiod of birds that were held under natural photoperiods at 27 degrees N for 2 years was similar to those who arrived the same year from their breeding grounds ( approximately 40 degrees N), suggesting that the experience of higher amplitude day-night (light-dark, LD) cycles during migratory and breeding seasons were not critical for the subsequent response (initiation-termination-reinitiation) cycle. Another experiment examined entrainment of the circadian photoperiodic rhythm in buntings by subjecting them to T=24+/-2 hr LD-cycles with 8 hr photophase and to T=22 and 24 hr with 11 hr photophase. The results showed a reduction in critical day length under T=22 hr LD-cycle. In the last experiment, we constructed an action spectrum for photoperiodic induction by exposing birds for 4.5 weeks to 13L:11D of white (control), blue (450 nm), or red (640 nm) light at irradiances ranging from 0.028 to 1.4 W m(-2). The threshold light irradiance for photoinduction was about 10-fold higher for blue light, than for red and white lights. These results conclude that the daily light of the environment regulates the endogenous program that times seasonal responses in body fattening and testicular cycles of the redheaded bunting.     

Diurnal changes in maize leaf photosynthesis : I. Carbon exchange rate, assimilate export rate, and enzyme activities

Diurnal changes in photosynthetic parameters and enzyme activities were characterized in greenhouse grown maize plants (Zea mays L. cv Pioneer 3184). Rates of net photosynthesis and assimilate export were highest at midday, coincident with maximum irradiance. During the day, assimilate export accounted for about 80% of net carbon fixation, and the maximum export rate (35 milligrams CH₂O per square decimeter per hour) was substantially higher than the relatively constant rate maintained through the night (5 milligrams CH₂O per square decimeter per hour). Activities of sucrose phosphate synthase and NADP-malate dehydrogenase showed pronounced diurnal fluctuations; maximum enzyme activities were generally coincident with highest light intensity. Reciprocal light/dark transfers of plants throughout the diurnal cycle revealed that both enzymes were deactivated by 30 minutes of darkness during the day, and they could both be substantially activated by 30 minutes of illumination at night. During 24 hours of extended darkness, sucrose phosphate synthase activity declined progressively to an almost undetectable level, but was activated after 1.5 hours of illumination. Thus, the diurnal fluctuation in maize sucrose phosphate synthase can be explained by some form of light modulation of enzyme activity and is not due to an endogenous rhythm in activity. No diurnal fluctuations were observed in the activities of NADP-malic enzyme or fructose 6-phosphate-2-kinase. Phosphoenolpyruvate carboxylase was activated by light to some extent (about 50%) when activity was measured under suboptimal conditions in vitro. The results suggested that the rates of sucrose formation and assimilate export were closely aligned with the rate of carbon fixation and the activation state of sucrose phosphate synthase.     

Germination of seeds in Jussiaea suffruticosa

Seeds of Jussiaea suffruticosa reach high germination percentagesonly when exposed to long periods of continuous illumination.The light reaction may be repeatedly reversed by short exposuresto red and far red light, thus being mediated by the phytochromesystem. Seeds also germinate at high percentages if exposedto various cycles of 1 hr light and 24 hr of darkness at 20°C.If the temperature in the periods of darkness is raised up to30°C or lowered to 10°C the promotive effect of lightis inhibited. High temperatures (35°C) during imbibitionhave a promotive effect, whereas a pure O₂ atmosphere decreasesthe response to light. KNO₃ and kinetin enhance the responseto light but do not provoke germination in the dark. Only ifseed coats are punctured or removed does germination in thedark occur. (Received January 14, 1969; )     

Phase relations between the circadian leaf movement rhythm and its zeitgeber within the range of entrainment in the cotton plant,Gossypium hirsutum L.

Abstract

The leaf movement rhythm of Gossypium hirsutum L. (cv. Lakshmi) could be entrained to 24 h LD cycles with different photofractions varying from 4 to 20 h such that the night peak position of the rhythm occurred during darkness. The phase angle (ψ) of the rhythm varied in a regular manner with different photoperiods of a 24 h LD cycle. Under 24 h LD cycles with different photoperiods, the leaf movement shows probable evidences for the concurrent participation of a ‘light‐on’ and ‘light‐off rhythm.     

The effects of blue and far red light on rhythmic leaflet movements in samanea and albizzia

The opening of excised Samanea saman pulvini is promoted by prolonged blue or far-red irradiation. Far-red effects are attributed partially but not completely to lowering of the Pfr level. Two hours of continuous or pulsed blue light or pulsed far-red light (total dosage = 2.2 × 10¹⁸ quanta per square centimeter in all cases) also phase shifts the rhythm in Samanea while two hours of continuous blue light phase shifts the rhythm in the related plant Albizzia julibrissin. The same pigments appear to regulate opening and rhythmic phase shifting. The blue light-induced phase response curve has smaller advance and delay peaks and differs in shape from the curve induced by brief red light pulses absorbed by phytochrome. The blue absorbing pigment has not been identified, but it does not appear to be phytochrome acting in a photoreversible mode.