The amplitudes of unit events in Limulus photoreceptors are modulated from an input that resembles the overall response

Light adaptation is a gain-control process that endows photoreceptors with large dynamic range. In invertebrates, this process appears to be mediated by a negative feedback that sets the amplitude of the isolated photon responses (bumps) by modulating an enzyme's rate of catalysis. This paper reports measurements of the feedback dynamics of Limulus from the responses to small modulations in light intensity. The responses show a noise that apparently arises from the random arrival of photons. We use a dynamic noiseanalysis technique to extract the cells's frequencyresponse transfer function for bump amplitude. Its ratio to the transfer function for the summed response of the cell has a simple form at low frequencies. This indicates that the origin of the feedback responsible for the adaptation is at a stage temporally close to the final conductance response. Moreover, the form of the transfer function suggests feedback by a chemical agent which is removed by a single enzymatic-like stage at low light intensity and by several such stages in parallel but with a spread of time constants at high intensity.This work was supported by grants from the Binational Science Foundation (BSF) Jerusalem, Israel and the Israel Academy of Sciences and Humanities, by NIH grant EY 1428, and by NSF grant DMS 8505442     

Fly photoreceptors and temperature: Relative UV-sensitivity is increased by cooling

Intracellular responses from blowfly photoreceptor cells were recorded at various temperatures in order to study the behaviour of the transduction system, with particular reference to spectral sensitivity. with decreased temperature the V-log I functions showed a reduction in amplitude and the responses showed a slowed time course. For double peaked spectral sensitivity function the UV or 350 nm peak was much less dependent on temperature than the peak in the visible region. The higher UV-sensitivity is interpreted in terms of the sensitizing pigment theory to indicate changes in the effectiveness of energy transfer between the two chromophores.     

A histochemical study of chondroid tissue in Limulus and Octopus

Summary In a comparative histochemical study of Octopus and Limulus chondroid tissues and mouse tracheal cartilage, it was demonstrated that both invertebrate chondroids behave as less acid mucopolysaccharides than those in mouse cartilage. Octopus chondroid was less reactive than Limulus chondroid. Cetyl pyridinium chloride blockage of toluidin blue metachramasia could be unblocked in mouse cartilage by an hour's treatment in 0.5 M KCl, but even extended periods in 2.0 M KCl failed to accomplish this in the invertebrate chondroids. With a number of methods for demonstrating proteins, Octopus chondroid was less reactive than Limulus chondroid. Limulus chondroid matrix was intensely stained by methods for demonstrating protein thiol groups, but essentially no staining was observed in Octopus chondroid matrix. These studies indicated that the composition and/or structure of matrix material in the invertebrate chondroids differ from one another, and in turn differ from conditions in vertebrate hyaline cartilage.This work was supported by a grant (HD-1499-04) and a Career Development Award (5-K3-6176-04) from the National Institute of Child Health and Human Development of the U.S. Public Health Service.A contribution of the Sea Horse Key Marine Laboratory of the University of Florida.     

Causal transfer function analysis to describe closed loop interactions between cardiovascular and cardiorespiratory variability signals

Although the concept of transfer function is intrinsically related to an input–output relationship, the traditional and widely used estimation method merges both feedback and feedforward interactions between the two analyzed signals. This limitation may endanger the reliability of transfer function analysis in biological systems characterized by closed loop interactions. In this study, a method for estimating the transfer function between closed loop interacting signals was proposed and validated in the field of cardiovascular and cardiorespiratory variability. The two analyzed signals x and y were described by a bivariate autoregressive model, and the causal transfer function from x to y was estimated after imposing causality by setting to zero the model coefficients representative of the reverse effects from y to x. The method was tested in simulations reproducing linear open and closed loop interactions, showing a better adherence of the causal transfer function to the theoretical curves with respect to the traditional approach in presence of non-negligible reverse effects. It was then applied in ten healthy young subjects to characterize the transfer functions from respiration to heart period (RR interval) and to systolic arterial pressure (SAP), and from SAP to RR interval. In the first two cases, the causal and non-causal transfer function estimates were comparable, indicating that respiration, acting as exogenous signal, sets an open loop relationship upon SAP and RR interval. On the contrary, causal and traditional transfer functions from SAP to RR were significantly different, suggesting the presence of a considerable influence on the opposite causal direction. Thus, the proposed causal approach seems to be appropriate for the estimation of parameters, like the gain and the phase lag from SAP to RR interval, which have a large clinical and physiological relevance.     

A quantitative description of the dynamics of excitation and inhibition in the eye of Limulus

By means of intracellular microelectrode techniques, we have measured the dynamics of the several processes which translate light stimulation into spike activity in the Limulus eye. The transductions from light to voltage and from voltage to spike rate, and the lateral inhibitory transduction from spike rate to voltage, we have characterized by transfer functions. We have checked the appropriateness of treating the eye as a system of linear transducers under our experimental conditions. The response of the eye to a large spot of light undergoing sine flicker has been correctly predicted.     

Cellular Mechanisms of Entrainment

This review summarizes our current understanding of the signal transduction cascade by which light causes phase shifts of the circadian oscillators found in the eye of Bulla and Aplysia. The isolated retina of these marine mollusks contains a circadian oscillator, a photoreceptor, and a light transduction pathway sufficient for entrainment. This preparation offers unique advantages for the cellular analysis of entrainment and the generation of circadian oscillations. There is evidence that similar cellular mechanisms may underlie mammalian and molluskan circadian oscillations. Thus, the models developed to explain entrainment in the molluskan retina are likely to have utility in exploring the mammalian supra-chiasmatic nucleus.     

Comparison of protein structure and genomic structure of human, rabbit, and limulus C-reactive proteins: Possible implications for function and evolution

The primary structures of human, rabbit, and Limulus C-reactive proteins (CRPs) have been compared by a computer program. Based on these data, a PAMs matrix (accepted point mutation per 100 residues) was constructed to generate topologies for the three proteins. Five trees with the shortest absolute length were generated, but only one positive tree was found. Using the relatively well-established distance between human and rabbit of 150 million years, we calculate that human and Limulus CRPs diverged at least 500 million years ago. The data indicate that the amino acid sequence indentity between Limulus CRPs and their mammalian counterparts is about 25%, strongly suggesting that human CRP, rabbit CRP, and Limulus CRPs share common ancestral genes. There are two highly conserved regions in the primary structures among the CRPs. Residues 52–67 in Limulus CRP and residues 51–66 in human CRP show identity in 10 of 16 positions, with 3 additional conservative replacements. This region of the molecule is thought to be involved in the binding of phosphorylcholine ligand. Residues 139–153 in Limulus CRP and residues 133–147 in human CRP show identity in 9 of 15 positions, with 5 additional conservative replacements. The biological function of this stretch of amino acid sequence is thought to be associated with the CA²⁺ binding of the CRPs.This article was presented during the proceedings of the International Conference on Macromolecular Structure and Function, held at the National Defence Medical College, Tokorozawa, Japan, December 1985.     

Two components of the receptor current are developed from distinct elementary signals in Limulus ventral nerve photoreceptor

Transient elementary currents, bumps, stimulated by short dim light flashes were measured in ventral nerve photoreceptors of Limulus. It is demonstrated that light activates two types of bumps, which form two distinct components of the receptor current at higher light intensities. The two bump types, which are both assumed to be activated by single absorbed photons, differ in current amplitude and kinetic parameters. The current amplitude of one bump type is smaller than 0.3 nA and that of the other type is in the usual current range of up to several nanoamperes. The average latency of small bumps measured from the short stimulus flash is shorter than that of the large bumps. The small bumps have slower activation kinetics than the large bumps. It is demonstrated that with increasing flash intensity the small bumps overlap first and form a macroscopic current, on top of which the large bumps are superimposed. Results indicate that a single absorbed photon selectively activates only one kind of the enzyme cascades evoking one bump type. We conclude that the active meta conformation of a rhodopsin molecule selectively binds a specific type of G-protein, which is involved in the stimulation of one of the transduction cascades. The two bump types, which are the elements of two macroscopic current components support the previous assumption that light activates different transduction mechanisms in Limulus photoreceptors.     

Membrane maintenance and electrical properties of photoreceptors of wild-type andrpa (receptor potential absent) mutant blowflies (Calliphora erythrocephala)

Summary The maintenance of photoreceptor cell membranes in the blowfly was investigated in relation to the diurnal cycle, age, and therpa (receptor potential absent) phototransduction mutation. The effect of disturbed membrane assembly on the electrical membrane properties was examined using single-electrode discontinuous current-clamp techniques. In wild-type flies the cross-sectional dimensions of the rhabdomeres were markedly reduced with age, and the quantity of synthetic organelles decreased concurrently, whereas no correlation was found between the diurnal cycle and membrane turnover. Therpa mutation is thought to block the visual transduction cascade in photoreceptor cells and to lead to degeneration of the photoreceptor cell bodies. The volume of rhabdomeres decreased markedly inrpa mutants and the quantity of synthetic organelles was reduced significantly, indicating an imbalance between photoreceptive membrane renewal and degradation. Also, the plasma membrane underwent degenerative changes. The passive electrical properties of photoreceptor cells — resting membrane voltages and input resistances — were only slightly changed from those of wild-type flies, although the photoreceptive membrane did not depolarize in response to light. This indicates no apparent disturbance in the function of the ionic channels in these membranes. Taken together, these results suggest that the photoreceptor cells need a functional phototransduction cascade with its feedback controls to maintain continuous renewal of rhabdomeres, but that the plasma membrane maintains its normal electrochemical properties despite extreme morphological degeneration of photoreceptor cell.     

Die Verarbeitung stationärer optischer Nachrichten im Komplexauge von Limulus

Summary The functional properties of the processing of visual information by the complex eye of Limulus was studied. The spatial distribution of activity that results in the optic nerve when the Limulus eye is exposed to a stationary optical pattern depends upon the transfer characteristics of two subsystems: the dioptric apparatus and the nervous interactions comprising the lateral inhibition system. — The transfer characteristic of the dioptric apparatus is determined by the sensitivity distribution function of single ommatidia. This distribution was measured and found to be approximately of Gauss-function type. The sensitivity falls off to 1/e at a distance of one ommatidium; thus the visual fields of adjacent ommatidia strongly overlap. As a consequence of the overlap, amplitudes of the spatial Fourier components, of which the brightness distribution of the optical surround is made up, are more and more reduced with increasing frequency in the intensity distribution on the receptor mosaic. The amplitude of the spatial frequency 1/=0,25 ( in units of interommatidial distance) is reduced to half of the maximum value, which is attained at zero frequency. It is shown that the amplitude frequency characteristic of the sensitivity distribution function has no zeros, which means that no loss of optical information results from overlap of visual fields. Thus the resolving power of the dioptric apparatus is limited only by the number of receptors per unit area. — The transfer characteristic of the lateral inhibition system in the Limulus eye depends on the distribution of the inhibitory coefficients around the individual receptors. This distribution function was determined from excitatory responses in the optic nerve elicited by a spatial light intensity step function on the receptor mosaic. It is found that this distribution is also Gaussian in form, but decays to 1/e at a distance of eight to nine ommatidia along the major axis of the eye. The average value of the inhibitory coefficients between adjacent ommatidia was found to be 0,025. The amplitude frequency response of the inhibitory system is constant for high spatial frequencies down to 1/=0,1 while amplitudes of lower frequency sinusoids are reduced down to nearly half of the maximum value at frequency zero. The amplitude frequency characteristic of the inhibitory system ensures a one to one correspondence between the intensity distribution on the receptor mosaic and the excitation distribution in the optic nerve. The overall transfer characteristic of the eye is derived from the transfer characteristics of the dioptric apparatus and the inhibitory system. This characteristic is of bandpass type with a maximum amplitude response at a frequency of 1/=0,07. The overall transfer characteristic was independently confirmed in a separate experiment. The nature of the overall transfer characteristic shows that the inhibitory system does not exactly correct for the overlap of the visual fields of single ommatidia, which in principal the system could do if the distributions of inhibitory coefficients and ommatidia sensitivity were equal. The overall transfer characteristic of the Limulus eye garantees a one to one correspondence between patterns in the optical surround and excitation distributions in the optic nerve. — The average values of the inhibitory coefficients derived from these experiments are at least a factor ten smaller than those determined directly by other investigators. Possible explanations of this discrepency are discussed. — In a separate chapter the overall transfer characteristic for eyes submerged in water is described. It was found that this characteristic does not differ from that determined in air for the eye region which was investigated in the experiments. This result is explained by two properties of the eye which are dependent on the refractive index of the surround medium and whose influences cancel each other: the visual fields of ommatidia are reduced under water, while the divergence angles between the optical axes of adjacent ommatidia also diminish.

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This research was supported in part by the United States Air Force under Grant No. AF-EOAR-62-41 and monitored by the European Office, Office of Aerospace Research.     

Evidence for hemocyanin formation in the compound eye of Squilla mantis (Crustacea,Stomatopoda)

Summary In the retina and in the subretinal space of the compound eye of Squilla mantis a special kind of pigment cell is present. The crystalline inclusions of this cell have been identified as hemocyanin, as determined (i) by the dimensional congruence of the crystalline substructure with the dimensions of isolated, purified hemocyanin, and (ii) by the immunofluorescence reaction using anti-hemicyanin antibodies. The ultrastructure of these cells, their location and the presence of crystalline bodies in their cytoplasm suggest that they are sites of hemocyanin synthesis and homologous to the cyanocytes or cyanoblasts of Limulus.Supported by grant No. 3,012-0.76 of the Swiss National Science Foundation     

Neuromuscular modulation in Limulus by both octopamine and proctolin

Both octopamine and proctolin potentiate nerve-evoked skeletal muscle contractions in the horseshoe crab, Limulus. The threshold concentration for octopamine was 10^?9 to 10^?8M, while for proctolin it was 3 × 10^?9M. Norepinephrine and dopamine produced effects similar to octopamine but at higher thresholds; tyramine and serotonin were ineffective. Octopamine caused significant increases in amplitudes of excitatory postsynaptic potentials (epsps) of muscle fibers, but had little effect on muscle fiber input resistance or membrane potential. Also, octopamine did not affect depolarization of muscle fibers and subsequent contraction due to the direct action of exogenously applied glutamate. These results suggest that octopamine potentiates nerve-evoked contractions primarily by facilitating release of neuromuscular transmitter. At concentrations above 10^?7M, however, octopamine sometimes caused muscle spikes in response to motoneuron stimulation, a finding that suggests that octopamine may also have some postsynaptic action. Proctolin potentiated the muscle contractions evoked by glutamate but had little effect on glutamate-evoked muscle fiber depolarization, muscle fiber input resistance, or membrane potential. Thus, proctolin appears to act directly on skeletal muscle to enhance contractility. The proctolin-induced potentiations of contraction were sometimes accompanied by modest increases in epsp amplitude, so that unlike lobster skeletal and Limulus cardiac neuromuscular preparations, proctolin may have a secondary direct synaptic effect. Both octopamine and proctolin have been found in Limulus cardiac ganglion. This potential access to the hemolymph and the relatively low threshold concentrations needed for physiological action suggest that octopamine and proctolin could function as hormonal modulators of neuromuscular function in Limulus.     

Light-induced changes in cAMP levels in Limulus photoreceptors

Illumination reduces cAMP levels about 3-fold in $\text{Limulus}$ photoreceptors. Cyclic GMP levels are not significantly changed under identical conditions. In addition, the cAMP content in dark-adapted photoreceptors is about 4-fold the content of cGMP. It is proposed that cAMP may be involved in the regulation of the metabolism or function of photoreceptor systems which contain the photopigment in the transducing surface membrane.     

Oscillations and multiple steady states in a cyclic gene model with repression

In this paper we study the cyclic gene model with repression considered by H. T. Banks and J. M. Mahaffy. Roughly, the model describes a biochemical feedback loop consisting of an integer number G of single gene reaction sequences in series. The model leads to a system of functional differential equations. We show that there is a qualitative difference in the dynamics between even and odd G if the feedback repression is sufficiently large. For even G, multiple stable steady states can coexist while for odd G, periodic orbits exist.This research was supported in part by the Air Force Office of Scientific Research under Contract #AFOSR-84-0376 and by the US Army Research Office under Contract #DAAG29-84-K-0082