Quantum Relations of the Rat Electroretinogram

The rat retina is uniform and contains almost exclusively rods. Therefore the rat eye, when uniformly illuminated, produces a gross electroretinogram (ERG) which is simply related to the activity of the individual retinal sources of the ERG. Characteristics of ERG's are shown on an intensity scale of the average number of quanta absorbed per rod per stimulus flash obtained by direct accurate measurement of all quantities involved. An independent check on the accuracy of these measurements is applied to pigment-bleaching data reported by Dowling (1963). When ERG characteristics are placed on this scale it is found that: (a) The b-wave can usually be observed when fewer than one out of two hundred rods absorbs a quantum, the threshold being determined by the noise of the preparation. (b) Near threshold the b-wave amplitude is proportional to intensity. (c) The a-wave appears when there are more than two to four absorptions per rod per flash. (d) The b-wave latency decreases with intensity, and the amplitude becomes proportional to the logarithm of intensity when fewer than one out of ten rods absorbs a quantum. This implies that the b-wave sources must combine excitation from more than one rod (probably more than seven). Therefore the b-wave cannot arise from independent rods or rod-bipolar synapses, but probably reflects activity of entire inner nuclear layer cells.     

THE LIGHT GROWTH RESPONSE AND THE GROWTH SYSTEM OF PHYCOMYCES

1. The Roscoe-Bunsen law holds for the light growth response of Phycomyces if the time component of stimulation is short. With exposures longer than a few seconds, the reaction time to light is determined by the intensity and not by the energy of the flash. 2. The possible nature of the very long latency in the response to light is considered in terms of the structure of the cell and its mechanism of growth. It is suggested that during the latency some substance produced by light in the protoplasm is transported centrifugally to the cell wall or outermost layer of protoplasm. 3. The total elongation occurring over a period of 1 to 2 hours is independent of flashes of light or temporary darkening. Light acts by facilitating some change already under way in the growth system, and during the principal phase of elongation is not a necessary or limiting factor for growth. 4. Judged by the reaction time, the original sensitivity is restored in the light system following exposure to light in about one-third the time required for equilibrium to be reattained in the growth system.     

The statistical detection of threshold signals in the retina

1. Photographic records of impulses from single ganglion cells in the cat's retina were made while the retina was stimulated by flashes occurring once a second. Ten flashes at each of several intensities near threshold were used. 2. For the purpose of statistical analysis, the number of impulses (x) falling within a critical period following each flash was used as an index of the response. Histograms of x were plotted and used to calculate rates of transfer of information by the ganglion cell for the case of an ideal experiment, the yes-no choice, in which flashes of intensity I and blanks are to be distinguished. 3. The information rate increased (a) with increasing stimulus intensity and (b) with the number of identical flashes or blanks presented successively in a block. The intensity chosen as threshold by the experimenter, who observed the impulses visually and aurally, corresponded to an average information rate for single flashes of 0.7 bit/flash, compared to the maximum possible rate of 1 bit/flash. A threshold intensity giving 0.4 or more bit/flash, if presented in blocks of six identical flashes, corresponded to 0.95 or more bit/block, or near certainty. Thus the calculation of information rates using the index x provides an estimate of threshold at least as sensitive as those obtained during an experiment, which were made only after observing the responses to five to ten flashes of the same intensity. 4. The index x has statistical properties similar to those of the "index of neural activity" used by Tanner and Swets (1954) in their statistical model of human vision, and represents a possible physical interpretation of their index. However, x gave values (0.5 to 1.5) of the parameter called the slope which were consistently smaller than their values (2.1 to 3.1).     

Flash kinetics and light intensity dependence of oxygen evolution in the blue-green alga Anacystis nidulans

Patterns of oxygen evolution in flashing light for the blue-green alga Anacystis nidulans are compared with those for broken spinach chloroplasts and whole cells of the green alga Chlorella pyrenoidosa. The oscillations of oxygen yield with flash number that occur in both Anacystis and Chlorella, display a greater degree of damping than do those of isolated spinach chloroplasts. The increase in damping results from a two- to threefold increase in the fraction (α) of reaction centers “missed” by a flash. The increase in α cannot be explained by non-saturating flash intensities or by the dark reduction of the oxidized intermediates formed by the flash. Anaerobic conditions markedly increase α in Anacystis and Chlorella but have no effect on α in broken spinach chloroplasts. The results signify that the mechanism of charge separation and water oxidation involved in all three organisms is the same, but that the pool of secondary electron acceptors between Photosystem II and Photosystem I is more reduced in the dark, in the algal cells, than in the isolated spinach chloroplasts.Oxygen evolution in flashing light for Anacystis and Chlorella show light saturation curves for the oxygen yield of the third flash (Y₃) that differ markedly from those of the steady-state flashes (Y_s). In experiments in which all flashes are uniformly attenuated, Y₃ requires nearly twice as much light as Y_s to reach half-saturation. Under these conditions Y₃ has a sigmoidal dependence on intensity, while that of Y_s is hyperbolic. These differences depend on the number of flashes attenuated. When any one of the first three flashes is attenuated, the variation of Y₃ with intensity resembles that of Y_s. When two of the first three flashes are attenuated, Y₃ is intermediate in shape between the two extremes. A quantitative interpretation of these results based on the model of Kok et al. (Kok, B., Forbush, B. and McGloin, M. (1970) Photochem. Photobiol. 11, 457–475, and Forbush, B., Kok, B. and McGloin, M. P. (1971) Photochem. Photobiol. 14, 307–321) fits the experimental data.     

Kinetics of the fluorescence yield of chlorophyll a2 in spinach chloroplasts at liquid nitrogen temperature du ring and following a 16 μs flash

If, at liquid nitrogen temperature, the initial fluorescence yield of chlorophyll a₂ is high (e.g. after preillumination), a 16 μs flash produces in a few microseconds a decrease in fluorescence yield, followed by an increase, which occurs after roughly 10–20 μs, when the intensity of the flash has become negligible. It is concluded that during a flash, a quencher or quenching state T is formed, which disappears in a dark reaction in a time of the order of 10 μs. The kinetics are the same and can be interpreted in the same way as the kinetics at room temperature earlier reported by Duysens et al. (Abstr. VI. Int. Congr. on Photobiol. Bochum 1972, No. 277).If the flash is given when the initial fluorescence yield is low, then the fluorescence yield increases only markedly at the end of the flash, when the intensity has become low. Even for a strong flash, the increase is only about 20% of the maximum increase attained after a large number of flashes. This indicates that at low temperature, in contradistinction to room temperature, the reduction of the primary oxidant Q is less efficient than the formation of the quencher T. For the interpretation of the experiments it was not necessary to introduce other light-induced quenchers than T, such as the oxidized primary reductant, P⁺.     

The induction kinetics of bacterial photophosphorylation. Threshold effects by the phosphate potential and correlation with the amplitude of the carotenoid absorption band shift

1. ATP synthesis (monitored by luciferin-luciferase) can be elicited by a single turnover flash of saturating intensity in chromatophores from Rhodopseudomonas capsulata, Kb1. The ATP yield from the first to the fourth turnover is strongly influenced by the phosphate potential: at high phosphate potential (?11.5 kcal/mol) no ATP is formed in the first three turnovers while at lower phosphate potential (?8.2 kcal/mol) the yield in the first flash is already one half of the maximum, which is reached after 2–3 turnovers.2. The response to ionophores indicates that the driving force for ATP synthesis in the first 20 turnovers is mainly given by a membrane potential. The amplitude of the carotenoid band shift shows that during a train of flashes an increasing ΔΨ is built up, which reaches a stationary level after a few turnovers; at high phosphate potential, therefore, more turnovers of the same photosynthetic unit are required to overcome an energetic threshold.3. After several (six to seven) flashes the ATP yield becomes constant, independently from the phosphate potential; the yield varies, however, as a function of dark time (t_d) between flashes, with an optimum for t_d = 160–320 ms.4. The decay kinetics of the high energy state generated by a long (125 ms) flash have been studied directly measuring the ATP yield produced in post-illumination by one single turnover flash, under conditions of phosphate potential (?10 kcal/mol), which will not allow ATP formation by one single turnover. The high energy state decays within 20 s after the illumination. The decay rate is strongly accelerated by 10^?8 M valinomycin.5. Under all the experimental conditions described, the amplitude of the carotenoid signal correlates univocally with the ATP yield per flash, demonstrating that this signal monitores accurately an energetic state of the membrane directly involved in ATP synthesis.6. Although values of the carotenoid signal much larger than the minimal threshold are present, relax slowly, and contribute to the energy input for phosphorylation, no ATP is formed unless electron flow is induced by a single turnover flash.7. The conclusions drawn are independent from the assumption that a ΔΨ between bulk phases is evaluable from the carotenoid signal.     

Transient Photoresponses of a Phototactic Microorganism, Haematococcus Pluvialis, Revealed by Light Scattering

A new method, using incoherent light scattering, has been developed to measure the flagellar beating frequency of swimming microorganisms. By means of this method, transient changes of flagellar beating frequency in response to white light flashes have been revealed in samples of a phototactic microorganism, Haematococcus pluvialis. An increase of flagellar beating frequency occurs when the flash dose (flash intensity × flash duration) is sufficient. Reciprocity between light intensity and flash duration holds for durations not exceeding 60-80 ms. For lower doses a bimodal distribution of flagellar beating frequency is revealed. No effect is observed for very low flashes or for red stimuli, whereas green light is effective. A detailed analysis of experimental results has allowed us to determine the characteristic time of the effect and follow its evolution. The correlation of this effect with visually observed behavior is discussed and a possible underlying mechanism is suggested.     

Probability of Occurrence of Discrete Potential Waves in the Eye of Limulus

Discrete potential waves can be recorded from cells in the eye of Limulus both in darkness and in dim illumination. With constant illumination the frequency of these waves is linearly related to light intensity and the distribution of intervals between waves follows an exponential function. The latency of waves evoked by short flashes of light is usually long and variable and the number of waves evoked by a flash varies randomly, obeying approximately a Poisson distribution. The results of experiments with flashes of light have been compared with the predictions derived from the hypotheses that one, two, or three quanta of light are required for production of one wave. The agreement of the data with the theory can be considered acceptable for the "one quantum" hypothesis, is less satisfactory for the "two quanta" hypothesis, and is very poor for the "three quanta" hypothesis.     

The initial response of Limulus ventral photoreceptors to bright flashes. Released calcium as a synergist to excitation

The leading edge of the response of Limulus ventral photoreceptors to brief flashes was investigated using a voltage clamp. The leading edge of responses increases linearly with flash intensity when dim flashes produce less than one photoisomerization per square micron of cell surface. Brighter flashes accelerate the initial portion of the response, resulting in a fourth-power relationship between the magnitude of the response at brief times after the flash and the flash intensity. The onset of this nonlinearity with increasing flash intensity is determined by the local density of photoisomerizations within the receptor. Responses to bright 10-15-mum-diam spots therefore rise faster than responses to diffuse flashes producing the same number of photoisomerizations within the receptor. Background illumination shortens the response latency and suppresses the initial nonlinearity. These phenomena can be explained by a model of transduction in which light activates two parallel cascades of reactions. Particles released by the first of these cascades open ionic channels, while the second produces an agent that accelerates the rate of production of particles by the first. Injection of the calcium buffer EGTA slows the initial portion of the response to bright flashes and suppresses its nonlinearity, which suggests that the accelerating agent released by the second cascade is calcium.     

Flash-induced photophosphorylation in Rhodospirillum rubrum chromatophores I. The relationship between cytochrome c-420 content and photophosphorylation

The content of cytochrome c-420 in Rhodospirillum rubrum chromatophores prepared by grinding with alumina is 5–10% of that in whole cells, and 20–40% in chromatophores by ‘French’ pressing.Flash-induced phosphorylation of various chromatophores which varied in cytochrome content from 7 to 40% is proportional to the cytochrome content. Extrapolating the cytochrome c-420 content to that observed in whole cells, a ratio $\text{ATP}\text{P}{}^{\mathrm{+}}\text{X}{}^{\mathrm{?}}$ near 1 is calculated. At low flash intensity the phosphorylation per flash is proportional to flash energy.Photophosphorylation in flashes given after a time of several minutes is only slightly dependent on the number of flashes. If the flashes are spaced from 0.1 to 10 s, relative phosphorylation in the first flash is about 70% and in the second 90% of that observed in the following flashes. Proton binding is not affected by the cytochrome c-420 content and a ratio of $\text{H}{}^{\mathrm{+}}\text{P}{}^{\mathrm{+}}\text{X}{}^{\mathrm{?}}$ of 2.3 was found.These results can be explained by a working hypothesis in which charge separation occurring at one reaction centre and the resulting electron transport mediated amongst others by c-420, results in the injection of two protons into an ATPase, this in contrast to a chemiosmotic mechanism, where the protons are released in the chromatophore inner space.     

Electron acceptors of bacterial photosynthetic reaction centers II. H+ binding coupled to secondary electron transfer in the quinone acceptor complex

The photoreduction of ubiquinone in the electron acceptor complex (Q₁Q₁₁) of photosynthetic reaction centers from Rhodopseudomonas sphaeroides, R26, was studied in a series of short, saturating flashes. The specific involvement of H⁺ in the reduction was revealed by the pH dependence of the electron transfer events and by net H⁺ binding during the formation of ubiquinol, which requires two turnovers of the photochemical act. On the first flash Q₁₁ receives an electron via Q₁ to form a stable ubisemiquinone anion (Q?^?₁₁); the second flash generates Q?^?₁. At low pH the two semiquinones rapidly disproportionate with the uptake of 2 H⁺, to produce Q₁₁H₂. This yields out-of-phase binary oscillations for the formation of anionic semiquinone and for H⁺ uptake. Above pH 6 there is a progressive increase in H⁺ binding on the first flash and an equivalent decrease in binding on the second flash until, at about pH 9.5, the extent of H⁺ binding is the same on all flashes. The semiquinone oscillations, however, are undiminished up to pH 9. It is suggested that a non-chromophoric, acid-base group undergoes a pK shift in response to the appearance of the anionic semiquinone and that this group is the site of protonation on the first flash. The acid-base group, which may be in the reaction center protein, appears to be subsequently involved in the protonation events leading to fully reduced ubiquinol. The other proton in the two electron reduction of ubiquinone is always taken up on the second flash and is bound directly to Q?^?₁₁. At pH values above 8.0, it is rate limiting for the disproportionation and the kinetics, which are diffusion controlled, are properly responsive to the prevailing pH. Below pH 8, however, a further step in the reaction mechanism was shown to be rate limiting for both H⁺ binding electron transfer following the second flash.     

Flash communication between the sexes of the firefly, Photuris lucicrescens

ABSTRACT. The courtship signal of the male firefly, Photuris lucicrescens Barber (Coleoptera, Lampyridae), is a brilliant crescendo flash which grows in intensity, reaches a peak and abruptly terminates. It was found to be triggered by a long neural burst from the brain. Males and females produce weak, twinkling flashes which induce male crescendo flashes. Female responses were triggered by a slowly rising intensity, and female response latency is therefore variable. Male and female P.lucicrescens fireflies produce two different types of flashes and both these flashes play an integral part in their courtship communication.     

On the O2 flash yields of two cyanophytes

We explored O₂ flash yield in two cyanophytes, Anacystis nidulans and Agmenellum quadruplicatum. On a rate-measuring electrode, a single flash gave a contour of O₂ evolution with a peak at about 10 ms which was maximum (100) for 680 nm background light. On 625 nm illumination the peak was smaller (62) but was followed by an increased tail of O₂ attributed to enhancement of the background. After a period of darkness, repetitive flashes (5 Hz) gave a highly damped initial oscillation in individual flash yields which finally reached steady state at 94% of the yield for 680 nm illumination. When O₂ of repetitive flashes was measured as an integrated flash yield the results was distinctive and similar to that for a continuous light 1 (680 nm). An apparent inhibition of respiration which persisted into the following dark period was taken as evidence for the Kok effect. With a concentration-measuring electrode, integrated flash yield vs. flash rate showed the same nonlinear behavior as O₂ rate vs. intensity of light 1. We draw three conclusions about the two cyanophytes. (a) The plastoquinone pool is substantially reduced in darkness. (b) Because of a high ratio of reaction centers, reaction center 1 / reaction center 2, for the two photoreactions, saturating flashes behave as light 1. (c) Because repetitive flashes are light 1, they also give a Kok effect which must be guarded against in measurements designed to count reaction centers.     

Photoinhibition by flash and continuous light of oxygen uptake by intact photosynthetic bacteria

The inhibition of respiration by continuous or flashing light has been studied in intact cells of different species of photosynthetic bacteria. For Rhodopseudomonas palustris, Rhodopseudomonas sphaeroides and Rhodopseudomonas capsulata, the inhibition by short actinic flashes shows a remarkable periodicity of two: each flash induces an inhibition of respiration, but a stimulation is observed after an even number of flashes. On the other hand, no oscillation is observed for Rhodospirillum rubrum and Rhodopseudomonas viridis cells. These different behaviours are explained by a difference in the redox state of the secondary electron acceptor as shown by the effect of ortho-phenanthroline on the amperometric signal. Addition of uncouplers (carbonyl cyanide m-chlorophenylhydrazone) or of an ATPase inhibitor (tri-N-butyl tin), has little effect on the oscillatory pattern induced by flash excitation. However, inhibition of respiration by continuous light is suppressed in the presence of carbonyl cyanide m-chlorophenylhydrazone. In the presence of tri-N-butyl tin the steady-state level is reached more rapidly than in the control experiment for a given light intensity. These results are interpreted as evidence of two modes of light inhibition of respiration in photosynthetic bacteria. A first type of inhibition, clearly shown under flash excitation, is due to interaction between respiratory and photosynthetic chains at the level of electron carriers. After each flash, an electron is diverted from the respiratory chain to the photooxidized reaction center. Because of the gating mechanism at the level of the secondary acceptor, the respiration is stimulated after an even number of flashes. The second mode of inhibition prevails under continuous illumination. Under these conditions, the rate of respiration is controlled essentially by the photoinduced proton electrochemical gradient.     

SPONTANEOUS AND STIMULATED BIOLUMINESCENCE OF THE DINOFLAGELLATE CERATOCORYS HORRZDA (PERIDINIALES)1

This is the first report of spontaneous bioluminescence in the autotrophic dinoflagellate Ceratocorys horrida von Stein. Bioluminescence was measured, using an automated data acquisition system, in a strain of cultured cells isolated from the Sargasso Sea. Ceratocorys horrida is only the second dinoflagellate species to exhibit rhythmicity in the rate of spontaneous flashing, flash quantum flux (intensity), and level of spontaneous glowing. The rate of spontaneous flashing was maximal during hours 2–4 of the dark phase [i.e. circadian time (CT)16–18 for a 14:10 h LD cycle (LD14:10)], with approximately 2% of the population flashing-min^?1, a rate approximately one order of magnitude greater than that of the dinoflagellate Gonyaulax polyedra. Flash quantum flux was also maximal during this period. Spontaneous flashes were 134 ms in duration with a maximum flux (intensity) of 3.1×10⁹ quanta-s^?1. Light emission presumably originated from blue fluorescent microsources distributed in the cell periphery and not from the spines. Values of both spontaneous flash rate and maximum flux were independent of cell concentration. Isolated cells also produced spontaneous flashes. Spontaneous glowing was dim except for a peak of 6.4× 10⁴quanta-s^?1 cell^?1, which occurred at CT22.9 for LD14:10 and at CT22.8 for LD12:12. The total integrated emission of spontaneous flashing and glowing during the dark phase was 4×10⁹ quantacell^?1, equivalent to the total stimulable luminescence. The rhythms for C. horrida flash and glow behavior were similar to those of Gonyaulax polyedra, although flash rate and quantum flux were greater. Spontaneous bioluminescence in C. horrida may be a circadian rhythm because it persisted for at least three cycles in constant dark conditions. This is also the first detailed study of the stimulated bioluminescence of C. horrida, which also displayed a diurnal rhythm. Cultures exhibited >200 times more mechanically stimulated bioluminescence during the dark phase than during the light phase. Mechanical stimulation during the dark phase resulted in 6.7 flashes. cell^?1; flashes were brighter and longer in duration than spontaneous flashes. Cruise-collected cells exhibited variability in quantum flux with few differences in flash kinetics. The role of dinoflagellate spontaneous bioluminescence in the dynamics of near-surface oceanic communities is unknown, but it may be an important source of natural in situ bioluminescence.     

The Rat Electroretinogram : I. Contrasting effects of adaptation on the amplitude and latency of the b-wave

Characteristics of the electroretinogram (ERG) produced by the essentially all rod eye of the rat are presented as functions of the number of quanta absorbed by each rod per stimulus flash. The ERG's were obtained with 1.5 msec. stimulus flashes and uniform illumination of the entire retina. Under these conditions, distortions in the ERG due to stray light are minimized, and the ERG more accurately reflects the activity of its retinal sources. The effects of background light and two forms of dark adaptation were studied and compared. The results, especially for the b-wave, permit an interpretation in terms of two distinct processes. One process appears to determine the b-wave latency. This process is almost independent of the state of adaptation of the retina. The other process does not affect the latency, but determines the b-wave threshold and amplitude. This process strongly depends upon the state of adaptation. Moreover, the effects of dark adaptation on this amplitude-determining process are almost identical with the effects of background light.     

Delayed fluorescence from Rhodopseudomonas sphaeroides following single flashes

Delayed fluorescence from Rhodopseudomonas sphaeroides chromatophores was studied with the use of short flashes for excitation. Although the delayed fluorescence probably arises from a back-reaction between the oxidized reaction center bacteriochlorophyll complex (P⁺) and the reduced electron acceptor (X^?), the decay of delayed fluorescence after a flash is much faster ($\text{τ}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{≈ 120 μ}\text{s}$) than the decay of P⁺X^?. The rapid decay of delayed fluorescence is not due to the uptake of a proton from the solution, nor to a change in membrane potential. It correlates with small optical absorbance changes at 450 and 770 nm which could reflect a change in the state of X^?.The intensity of the delayed fluorescence is 11–18-fold greater if the excitation flashes are spaced 2 s apart than it is if they are 30 s apart. The enhancement of delayed fluorescence at high flash repetition rates occurs only at redox potentials which are low enough (< + 240 mV) so that electron donors are available to reduce P⁺X^? to PX^? in part of the reaction center population. The enhancement decays between flashes as PX^? is reoxidized to PX, as measured by the recovery of photochemical activity. Evidently, the reduction of P⁺X^? to PX^? leads to the storage of free energy that can be used on a subsequent flash to promote delayed fluorescence. The reduction of P⁺X^? also is associated with a carotenoid spectral shift which decays as PX^? is reoxidized to PX. Although this suggests that the free energy which supports the delayed fluorescence might be stored as a membrane potential, the ionophore gramicidin D only partially inhibits the enhancement of delayed fluorescence. With widely separated flashes, gramicidin has no effect on delayed fluorescence.At redox potentials low enough to keep X fully reduced, delayed fluorescence of the type described above does not occur, but one can detect weak luminescence which probably is due to phosphorescence of a protoporphyrin.     

Response entrainment of movement fibers in the optic tract of crayfish

The response properties of jittery movement fibers (JMF) in the crayfish optic tract reacting to a non-moving temporally patterned light were analyzed. The JMFs usually show no response during the regular flickering of stationary light with a flash duration of less than 50 msec when the stimulus frequency is between 4 and 20 per second; however they do respond when the flickering stops if a certain number of flashes have been given. The response appears about 50 msec after the first missing flash, i.e., the latency of the response after the last flash of the train changed from 100 to 300 msec. Thus, the “off” response at the end of the flicker is entrained to the stimulus repetition interval and locked onto the time of the first missing flash. The response of a sustaining fiber to an identical stimulus has quite different features as illustrated in Fig. 2. Some of the fibers show responses to the beginning part of the flicker but not necessarily to each flash, and habituate after several flashes. When a single flash longer than 250 msec is given, the fiber shows an “off” response with about 50 msec latency, as it does to sustained light. Some fibers show a double burst of “off” discharge to single flashes; the first at 50 msec is followed after 120 msec by the second one. However, when the flash duration is between 250 and 50 msec, a single flash elicits little or no response. The latency of the “off” response is as much as 300 msec for short single flashes less than 50 msec. An “on” response to flashes of light is observed when the inter-stimulus interval is more than 5 sec. The responses to the beginning part of flicker train are not simply locked to the just preceding flash except the “on” response to the very first one, but they can be the long latency responses to the flash before that. This response is modified in latency by the succeeding flashes in flicker trains and becomes entrained to the missing flash. Four types of entrainment are classified on the basis of the change in latency from the missing flash with regard to the number of flashes in a train. In most cases, 10 flashes are sufficient to entrain the response to the first missing flash. Non-resposiveness, i.e., habituation, during a regular flicker, may be due to an active inhibitory process, initiated by each succeeding light pulse. The response to the missing flash, therefore results from a disinhibited modified response to the last flash. Some JMFs continue to respond to the flicker even after a considerable number of flashes but only when the repetition interval is about 120 msec corresponding well to the interval of the double burst “off” discharge, thus the JMF has a resonant frequency of about 8 Hz. The JMFs appear to be acting as an irregularity detector in temporal sequence.     

Reactions between primary and secondary acceptors of Photosystem II in Chlorella Pyrenoidosa under anaerobic conditions as studied by chlorophyll a fluorescence

The kinetics of the fluorescence yield Ф of chlorophyll a in Chlorella pyrenoidosa were studied under anaerobic conditions in the time range from 50 μs to several minutes after short ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}\text{= 30}\text{ns}$ or 5 μs) saturating flashes. The fluorescence yield “in the dark” increased from $\text{Ф = 1}$ at the beginning to $\text{Ф ≈ 5}$ in about 3 h when single flashes separated by dark intervals of about 3 min were given.After one saturating flash, Ф increased to a maximum value (4–5) at 50 μs, then Ф decreased to about 3 with a half time of about 10 ms and to the initial value with a half time of about 2 s. When two flashes separated by 0.2 s were given, the first phase of the decrease after the second flash occurred within 2 ms. After one flash given at high initial fluorescence yield, the 10-ms decay was followed by a 10 s increase to the initial value. After the two flashes 0.2 s apart, the rapid decay was not follewed by a slow increase.These and other experiments provided additional evidence for and extend an earlier hypothesis concerning the acceptor complex of Photosystem II (Bouges-Bocquet, B. (1973) Biochim. Biophys. Acta 314, 250–256; Velthuys, B. R. and Amesz, J. (1974) Biochim. Biophys. Acta 333, 85–94): reaction center 2 contains an acceptor complex QR consisting of an electron-transferring primary acceptor molecule Q, and a secondary electron acceptor R, which can accept two electrons in succession, but transfers two electrons simultaneously to a molecule of the tertiary acceptor pool, containing plastoquinone (A). Furthermore, the kinetics indicate that 2 reactions centers of System I, excited by a short flash, cooperate directly or indirectly in oxidizing a plastohydroquinone molecule (A^2?). If initially all components between photoreaction 1 and 2 are in the reduced state the following sequence of reactions occurs after a flash has oxidised A^2? via System I: Q^?R^2? + A → Q^?R + A^2? → QR^? + A^2?. During anaerobiosis two slow reactions manifest themselves: the reduction of R (and A) within 1 s, presumably by an endogenous electron donor D₁, and the reduction of Q in about 10 s when R is in the state R^? and A in the state A^2?. An endogenous electron donor, D₂, and Q^? compete in reducing the photooxidized donor complex of System II in reactions with half times of the order of 1 s.