Light and dark adaptation in Phycomyces phototropism

Light and dark adaptation of the phototropism of Phycomyces sporangiophores were analyzed in the intensity range of 10(-7)-6 W X m- 2. The experiments were designed to test the validity of the Delbruck- Reichardt model of adaptation (Delbruck, M., and W. Reichardt, 1956, Cellular Mechanisms in Differentiation and Growth, 3-44), and the kinetics were measured by the phototropic delay method. We found that their model describes adequately only changes of the adaptation level after small, relatively short intensity changes. For dark adaptation, we found a biphasic decay with two time constants of b1 = 1-2 min and b2 = 6.5-10 min. The model fails for light adaptation, in which the level of adaptation can overshoot the actual intensity level before it relaxes to the new intensity. The light adaptation kinetics depend critically on the height of the applied pulse as well as the intensity range. Both these features are incompatible with the Delbruck-Reichardt model and indicate that light and dark adaptation are regulated by different mechanisms. The comparison of the dark adaptation kinetics with the time course of the dark growth response shows that Phycomyces has two adaptation mechanisms: an input adaptation, which operates for the range adjustment, and an output adaptation, which directly modulates the growth response. The analysis of four different types of behavioral mutants permitted a partial genetic dissection of the adaptation mechanism. The hypertropic strain L82 and mutants with defects in the madA gene have qualitatively the same adaptation behavior as the wild type; however, the adaptation constants are altered in these strains. Mutation of the madB gene leads to loss of the fast component of the dark adaptation kinetics and to overshooting of the light adaptation under conditions where the wild type does not overshoot. Another mutant with a defect in the madC gene shows abnormal behavior after steps up in light intensity. Since the madB and madC mutants have been associated with the receptor pigment, we infer that at least part of the adaptation process is mediated by the receptor pigment.     

Wavelength dependence of dark adaptation in Phycomyces phototropism

The wavelength dependence of phototropic dark adaptation in Phycomyces was studied between 347 and 545 nm. Dark adaptation kinetics were measured for wavelengths of 383, 409, 477, and 507 nm in the intensity range from 6.2 X 10(-2) to 2 X 10(-7) W X m-2. At these wavelengths, dark adaptation follows a biexponential decay as found previously with broadband blue light (Russo, V. E. A., and P. Galland, 1980, Struct. Bonding., 41:71; Lipson, E. D., and S. M. Block, 1983, J. Gen. Physiol., 81:845). We have found that the time constants of the fast and slow components depend critically on the wavelength. At 507 nm, dark adaptation kinetics were found to be monophasic. The phototropic latency after a step down by a factor of 500 was measured for 19 different wavelengths. Maximal latencies were found at 383, 477, and 530 nm; minimal latencies were found at 409 and 507 nm. With irradiation programs that employ different wavelengths before and after the step down, the dark adaptation kinetics depend critically on the sequence in which the two wavelengths are given. We have found too that not only do the adaptation kinetics vary with wavelength, but so also do the phototropic bending rate and the phototropic latencies in experiments without intensity change. The results imply that more than one photoreceptor is mediating phototropism in Phycomyces and that sensory adaptation is regulated by these photoreceptors.     

White noise analysis of Phycomyces light growth response system. II. Extended intensity ranges.

By means of white gaussian noise stimulation, the Wiener kernels are derived for the Phycomyces light growth response for a variety of intensity conditions. In one experiment the intensity I, rather than log I, is used as the input variable. Under the very limited dynamic range of that experiment, the response is fairly linear. To examine the dependence of the kernels on dynamic range, a series of experiments were performed in which the range of log I was halved and doubled relative to normal. The amplitude of the kernels, but not the time course, is affected strongly by the choice of dynamic range, and the dependence reveals large-scale nonlinearities not evident in the kernels themselves. In addition kernels are evaluated for experiments at a number of absolute intensity levels ranging from 10(-12) to 10(-3) W/cm2. The kernel amplitudes are maximal at about 10(-6) W/cm2. At 10(-12) W/cm2, just above the absolute threshold, the respond is very small. The falloff at high intensity, attributable to inactivation of the photoreceptor, is analyzed in the framework of a first-order pigment kinetics model, yielding estimates for the partial extinction coefficient for inactivation epsilonI455 = (1.5 +/- 0.2) X 10(4) liter/mol-cm and a regeneration time constant of tau = (2.7 +/- 0.6) min. A model is introduced which associates the processes of adaptation and photoreceptor inactivation. The model predicts that the time constants for adaptation and pigment should be identical. This prediction is consistent with values in this and the preceding paper. The effects of pigment inactivation are simulated by a linear electronic analog circuit element, which may be cascaded with the linear simulator circuit in the preceding paper.     

Threshold and adaptation in Phycomyces. Their interrelation and regulation by light

The absolute light sensitivity of Phycomyces sporangiophores was determined by analyzing the intensity dependence of the phototropic bending rate and of the light growth and dark growth responses to step changes of the intensity. We found that the different methods give approximately the same results for the wild-type strain, as well as for several behavioral mutants with defects in the genes madA, madB, and madC. A crucial factor in the determination of thresholds is the light intensity at which the strains grow during the 4 d after inoculation and prior to the experiment. When the wild-type strain grows in the dark, its threshold for the bending rate is 10(-9) W X m-2, compared with 2 X 10(-7) W X m-2 when it is grown under continuous illumination. Further, the maximal bending rate is twice as high in dark-grown strains. This phenomenon is further complicated by the fact that the diameter and growth rate of the sporangiophores also depend on the illumination conditions prior to the experiment: light-grown sporangiophores have an increased diameter and an increased growth rate compared with dark-grown ones. Some of the behavioral mutants, however, are indifferent to this form of light control. Another factor that is controlled by the growth conditions is adaptation: the kinetics of dark adaptation are slower in light-grown sporangiophores than in dark-grown ones. We found empirically a positive correlation between the slower dark adaptation constant and the threshold of the bending rate, which shows that the two underlying phenomena are functionally related.     

White noise analysis of Phycomyces light growth response system. I. Normal intensity range.

The Wiener-Lee-Schetzen method for the identification of a nonlinear system through white gaussian noise stimulation was applied to the transient light growth response of the sporangiophore of Phycomyces. In order to cover a moderate dynamic range of light intensity I, the imput variable was defined to be log I. The experiments were performed in the normal range of light intensity, centered about I0 = 10(-6) W/cm2. The kernels of the Wierner functionals were computed up to second order. Within the range of a few decades the system is reasonably linear with log I. The main nonlinear feature of the second-order kernel corresponds to the property of rectification. Power spectral analysis reveals that the slow dynamics of the system are of at least fifth order. The system can be represented approximately by a linear transfer function, including a first-order high-pass (adaptation) filter with a 4 min time constant and an underdamped fourth-order low-pass filter. Accordingly a linear electronic circuit was constructed to simulate the small scale response characteristics. In terms of the adaptation model of Delbrück and Reichardt (1956, in Cellular Mechanisms in Differentiation and Growth, Princeton University Press), kernels were deduced for the dynamic dependence of the growth velocity (output) on the "subjective intensity", a presumed internal variable. Finally the linear electronic simulator above was generalized to accommodate the large scale nonlinearity of the adaptation model and to serve as a tool for deeper test of the model.     

DARK ADAPTATION IN DINEUTES

The level of dark adaptation of the whirligig beetle can be measured in terms of the threshold intensity calling forth a response. The course of dark adaptation was determined at levels of light adaptation of 6.5, 91.6, and 6100 foot-candles. All data can be fitted by the same curve. This indicates that dark adaptation follows parts of the same course irrespective of the level of light adaptation. The intensity of the adapting light determines the level at which dark adaptation will begin. The relation between log aI ₀ (instantaneous threshold) and log of adapting light intensity is linear over the range studied.     

Increased intracellular sodium mimics some but not all aspects of photoreceptor adaptation in the ventral eye of Limulus

The effects of the intracellular iontophoretic injection of Na+ ions have been quantitatively compared with adaptation in ventral photoreceptors of Limulus. We find that: (a) both light adaptation and sodium injection are associated with a decrease in the variability of the threshold response amplitued; (b) both light adaptation and sodium injection are associated with a decrease in the absolute value of the temporal dispersion of the threshold response time delay; (c) the same template curve adequately fits the intensity response relationships measured under light adaptation and Na+ injection; (d) both light adaptation and Na+ injection produce a fourfold decrease in response time delay for a desensitization of 3 log units; (e) the time coures of light adaptation and dark adaptation is significantly faster than the onset of and recovery from desensitization produced by Na+ injection; (f) unlike local illumination, Na+ injection does not produce localized desensitization of the photoreceptor. These findings suggest that a rise in intracellular Na+ concentration makes at most only a minor contribution (probably less than 5%) to the total adaptation of these receptors in the intensity range we have examined (up to 3 log units above absolute threshold). However, changes in intracellular Na+ concentration may contribute to certain components of light and dark adaptation in these receptors.     

System analysis of Phycomyces light-growth response with sum-of-sinusoids test stimuli.

The light-growth response of Phycomyces has been studied with the sum-of-sinusoids method of nonlinear system identification (Victor, J.D., and R.M. Shapley, 1980, Biophys. J., 29:459). This transient response of the sporangiophore has been treated as a black-box system with one input (logarithm of the light intensity, I) and one output (elongation rate). The light intensity was modulated so that log I, as a function of time, was a sum of sinusoids. The log-mean intensity was 10(-4) W m-2 and the wavelength was 477 nm. The first- and second-order frequency kernels, which represent the linear and nonlinear behavior of the system, were obtained from the Fourier transform of the response at the appropriate component and combination frequencies. Although the first-order kernel accounts for most of the response, there remains a significant nonlinearity beyond the logarithmic transducer presumed to occur at the input of the sensory transduction chain. From the analysis of the frequency kernels, we have derived a dynamic nonlinear model of the light-growth response system. The model consists of a nonlinear subsystem followed by a linear subsystem. The model parameters were estimated from a combined nonlinear least-squares fit to the first- and second-order frequency kernels.     

Light and dark active phosphodiesterase regulation in salamander rods

We studied the activation of 3',5'-cyclic guanosine monophosphate (cGMP) phosphodiesterase (PDE) by using a cell-permeant enzyme inhibitor. Rods of Ambystoma tigrinum held in a suction electrode were jumped into a stream of 3-isobutyl-1-methylxanthine (IBMX), 0.01-1 mM. Initial transient light-sensitive currents fit the notion that dark and light-activated forms of PDE contributed independently to metabolic activity and were equivalently inhibited by IBMX (apparent Ki 30 microns). Inhibition developed within 50 ms, producing a step decrease of enzyme velocity, which could be offset by activation with flashes or steps of light. The dark PDE activity was equivalent to light activation of enzyme by 1,000 isomerization rod-1s-1, sufficient to hydrolyze the free cGMP pool (1/e) in 0.6 s. Steady light activated PDE in linear proportion to isomerization rate, the range from darkness to current saturation amounting to a 10-fold increase. The conditions for simultaneous onset of inhibitor and illumination to produce no net change of membrane current defined the apparent lifetime of light-activated PDE, TPDE* = 0.9 s, which was independent of both background illumination and current over the range 0-3 x 10(5) isomerization s-1, from 50 to 0 pA. Adaptation was a function of current rather than isomerization: jumps with different proportions of IBMX concentration to steady light intensity produced equal currents, and followed the same course of adaptation in maintained light, despite a 10-fold difference of illumination. Judged from the delay between IBMX- and light-induced currents, the dominant feedback regulatory site comes after PDE on the signal path. The dark active PDE affects the hydrolytic flux and cytoplasmic diffusion of cGMP, as well as the proportional range of the cGMP activity signal in response to light.     

Adapting-bump model for eccentric cells of Limulus

Light-evoked intracellular voltage noise records have been obtained from Limulus eccentric cells, from threshold light intensity to an intensity .10(5) times threshold. These data are analyzed in terms of a simple "adapting-bump" noise model. It is shown how the model yields a data reduction procedure that slightly generalizes the familiar use of Campbell's theorem for Poisson shot noise: the correlative effect of adaptation amends Campbell's theorem by a single multiplicative factor, which may be estimated directly from the power spectrum of the noise data. The model also permits direct estimation of the bump shape from the power spectrum. The bump shape estimated from noise at dim light is in excellent agreement with the average shape of bumps observed directly in the dark. The data yield a bump rate that is linear with light up through about 50 times threshold intensity but that falls short of linearity by a factor of 35 at the brightest light. The bump height decreases as the -0.4 power of light intensity across the entire range. Bump duration decreases by a factor of 2 across the entire range, and the adaptation correlation factor descends from unity to about one-third. The modest change of the adaptation correlation shows that naive application of Campbell's theorem to such data is adequate for rough estimation of the model's physiological parameters. This simple accounting for all the data gives support to the adapting-bump model.     

System analysis of Phycomyces light-growth response. Photoreceptor and hypertropic mutants.

The light-growth responses of Phycomyces behavioral mutants, defective in genes madB, madC, and madH, were studied with the sum-of-sinusoids method of system identification. Modified phototropic action spectra of these mutants have indicated that they have altered photoreceptors (P. Galland and E.D. Lipson, 1985, Photochem. Photobiol. 41:331). In the two preceding papers, a kinetic model of the light-growth response system was developed and applied to wild-type frequency kernels at several wavelengths and temperatures. The present mutant studies were conducted at wavelength 477 nm. The log-mean intensity was 6 X 10(-2)W m-2 for the madB and madC night-blind mutants, and 10(-4)W m-2 for the madH hypertropic mutant. The prolonged light-growth responses of the madB and madC mutants are reflected in the reduced dynamic order of their frequency kernels. The linear response of the hypertropic mutant is essentially normal, but its nonlinear behavior shows modified dynamics. The behavior of these mutants can be accounted for by suitable modifications of the parametric model of the system. These modifications together support the hypothesis that an integrated complex mediates sensory transduction in the light responses and other responses of the sporangiophore.     

Light adaptation in turtle cones. Testing and analysis of a model for phototransduction.

Light adaptation in cones was characterized by measuring the changes in temporal frequency responses to sinusoidal modulation of light around various mean levels spanning a range of four log units. We have shown previously that some aspects of cone adaptation behavior can be accounted for by a biochemical kinetic model for phototransduction in which adaptation is mediated largely by a sigmoidal dependence of guanylate cyclase activity on the concentration of free cytoplasmic Ca2+, ([Ca2+]i) (Sneyd and Tranchina, 1989). Here we extend the model by incorporating electrogenic Na+/K+ exchange, and the model is put to further tests by simulating experiments in the literature. It accounts for (a) speeding up of the impulse response, transition from monophasic to biphasic waveform, and improvement in contrast sensitivity with increasing background light level, I0; (b) linearity of the response to moderate modulations around I0; (c) shift of the intensity-response function (linear vs. log coordinates) with change in I0 (Normann and Perlman, 1979); the dark-adapted curve adheres closely to the Naka-Rushton equation; (d) steepening of the sensitivity vs. I0 function with [Ca2+]i fixed at its dark level, [Ca2+]i dark; (Matthews et al., 1988, 1990); (e) steepening of the steady-state intensity-response function when [Ca2+]i is held fixed at its dark level (Matthews et al., 1988; 1990); (f) shifting of a steep template saturation curve for normalized photocurrent vs. light-step intensity when the response is measured at fixed times and [Ca2+]i is held fixed at [Ca2+]i dark (Nakatani and Yau, 1988). Furthermore, the predicted dependence of guanylate cyclase activity on [Ca2+] closely matches a cooperative inhibition equation suggested by the experimental results of Koch and Stryer (1988) on cyclase activity in bovine rods. Finally, the model predicts that some changes in response kinetics with background light will still be present, even when [Ca2+]i is held fixed at [Ca2]i dark.     

THE COURSE OF ROD DARK ADAPTATION AS INFLUENCED BY THE INTENSITY AND DURATION OF PRE-ADAPTATION TO LIGHT

An increase in the degree of light adaptation causes a decrease in the slope of the subsequent rod dark adaptation function and a displacement of the function to the right on the time axis. Over a wide range, these changes occur to the same extent whether the increase in the degree of light adaptation is produced by raising the intensity or by prolonging the exposure. Within these limits, the Bunsen-Roscoe reciprocity law applies to the intensity and duration of pre-exposure. Over a still wider range, dark adaptation has the same course following brief exposure to a bright light as it has following prolonged exposure to a dim light, provided the degree of light adaptation is the same in both instances (as indicated by identical initial dark adaptation thresholds).     

Diminution and enlargement of the mosquito rhabdom in light and darkness

The rhabdoms of the larval ocelli of the mosquito Aedes aegypti undergo morphological light and dark adaptation over periods of hours. The rhabdom enlarges during dark adaptation and grows smaller during light adaptation. Diminution is exponential, enlargement linear, and rates of change are proportional to log light intensity. Rhabdoms maintained at a constant intensity level off at a constant volume proportional to log intensity. We argue that changes in rhabdom volume after changes in light intensity reflect an influence of light on the turnover of photoreceptro membrane, and that the volumes at which rhabdoms level off represent equilibria between opposed processes of membrane loss and renewal.     

DARK ADAPTATION AND THE DARK GROWTH RESPONSE OF PHYCOMYCES

Light-adapted sporangiophores of the fungus Phycomyces respond to sudden darkening by a temporary decrease in the rate of elongation, after a latent period of several minutes. The reaction time of this "dark growth" response is compound like that of the "light growth" response. It is, moreover, shorter the more intense the previous illumination. The rate of dark adaptation following adaptation to a very large range of light intensities is found to be proportional to the logarithm of the preceding light intensity. It is shown that a constant amount of dark adaptation takes place before the response occurs. On the assumption that changes in the rate of growth reflect changes in the concentration of a substance which at constant light intensity is in equilibrium with a light-sensitive material, possible equations for such a photostationary state are examined. The most reasonable formulation requires that the partial velocity of the "light" reaction be taken proportional to log I instead of to I directly.     

Photogravitropic equilibrium in Avena coleoptiles: fluence rate-response relationships and dependence on dark adaptation and clinostating

Phototropism of Avena coleoptiles was measured in response to blue-light irradiation lasting between 2 and 24 h. During this time the coleoptiles established a bending angle of photogravitropic equilibrium that was dependent on the time of irradiation and also on the pretreatment in light or darkness prior to stimulation. The absolute threshold for the photogravitropic equilibrium in response to blue light was 10(-8) micromol m(-2) s(-1). Photon fluence rate-response curves, which were generated after several hours of dark adaptation, had a characteristic shape with a prominent optimum in the middle of the dynamic range. Curves which were generated without prior dark adaptation displayed no such optimum. Clinostating dark-adapted coleoptiles caused an increase of sensitivity and responsiveness during a 2-h period of unilateral irradiation. The advantages and the drawbacks of long-term irradiation experiments for the investigation of phototropism and the generation of action spectra are discussed.     

Light Induced Concentration Changes of Adenosine-Triphosphate in Phycomyces Sporangiophores

The concentrations of extractable adenosine triphosphate (ATP) following the induction of positive light-growth responses in Phycomyces sporangiophores by blue light stimuli have been measured by means of the luciferin-luciferase assay. The ATP concentration in the light-sensitive growing zone increases 30 to 50% within 30 seconds after the start of a light stimulus and returns to the normal adapted level within 1 minute after stimulation. The ATP concentration is constant for any level of light adaptation and is uniform along the length of sporangiophores even though the light sensitivity is confined to a growing zone less than 5 mm long. These results suggest that one of the initial biochemical steps after a light stimulus is the production of extractable ATP.     

Comparative Studies on Dark Adaptation in the Compound Eyes of Nocturnal and Diurnal Lepidoptera

A comparative analysis has been carried out of the time course and range of dark adaptation in the compound eyes of some common butterflies and noctuid moths (Lepidoptera). The change in sensitivity of the eye during dark adaptation was determined by measurements of the intensity of illumination necessary to elicit an electrical response of a given magnitude of the eye. It was found that the curve for dark adaptation in the diurnal species was smooth. The range of adaptive change varied in different species but usually did not cover more than 1 to 1.5 log units. In the nocturnal species the dark adaptation was found to proceed in two phases. The first phase was usually completed in less than 10 minutes and covered a range of 1 to 1.5 log units. The second phase was more prolonged and covered a range of 2 to 3 log units. In some of the experiments on nocturnal species the second phase failed to appear. Measurements of the size of the response at different intensities showed that the intensity/amplitude relationship was the same in the light-adapted eye as in the dark-adapted eye. In the nocturnal insects the response of the eye in the light-adapted condition was about 20 per cent of that in the dark-adapted eye, while in diurnal insects it was about 60 per cent.     

The effect of light adaptation on the dynamic properties of phototransduction in the fly,Phormia regina

The static and dynamic characteristics of phototransduction were studied in photoreceptors of the compound eye of the fly Phormia regina (Calliphoridae) using a green light emitting diode driven by a controlled current source. The LED provides sufficiently intense light to investigate the behaviour of the receptors over about half of the dark adapted range of the response versus log intensity curve. The effects of constant adapting light intensities upon the step response and upon the frequency response and coherence functions were examined. Using both methods the effect of light adaptation upon receptor sensitivity can be closely approximated by a similar linear dependence of log sensitivity upon log adapting intensity. However, there was no reliably detectable effect of light adaptation upon the time constant of the response over the range of adapting intensities used.Abbreviation LED Light Emitting Diode     

Photoconversion from the light-adapted to the dark-adapted state of bacteriorhodopsin.

Dark and light adaptation of bacteriorhodopsin in purple membrane multilayers at less than 100% relative humidity differs from that seen in suspensions. Equilibrium between the two bacteriorhodopsin isomers (bR cis 550 and bR trans 570) in the light-adapted state becomes dependent on the wavelength of actinic light. Excitation at the red edge of the visible absorption band causes dark adaptation in a light-adapted sample. Using polarized actinic and measuring light, we show that acceleration of the dark adaptation through heating by actinic light cannot explain this observation. A light-driven bR trans 570 to bR cis 550 reaction that competes with the well-known 13 cis-to-all-trans light adaptation reaction must exist under our experimental conditions. Trans-to-cis conversion is a one-photon process distinct from the two photon process observed by others in purple membrane suspensions (Sperling, W., C. N. Rafferty, K. D. Kohl, and N. A. Dencher, 1978, FEBS (Fed. Eur. Biochem. Soc.) Lett. 97:129-132). Its quantum efficiency increases monotonously on reducing the hydration level, and is paralleled by an increase in the lifetime of the M410 intermediate of the trans photocycle. We suggest that at this point a branch leads from the all-trans into the 13-cis photocycle. It is probably the same reaction that causes the reduced light adaptation in monomeric bacteriorhodopsin (Casadio, R., H. Gutowitz, P. Mowery, M. Taylor, and W. Stoeckenius, 1980, Biochim. Biophys. Acta. 590:13-23; Casadio, R., and W. Stoeckenius, 1980, Biochemistry. 19:3374-3381).