Roger C. Hardie

The trp gene is essential for a light-activated Ca2+ channel in Drosophila photoreceptors

Invertebrate phototransduction is an important model system for studying the ubiquitous inositol-lipid signaling system. In the transient receptor potential (trp) mutant, one of the most intensively studied transduction mutants of Drosophila, the light response quickly declines to baseline during prolonged intense light. Using whole-cell recordings from Drosophila photoreceptors, we show that the wild-type response is mediated by at least two functionally distinct classes of light-sensitive channels and that both the trp mutation and a Ca2+ channel blocker (La3+) selectively abolish one class of channel with high Ca2+ permeability. Evidence is also presented that Ca2+ is necessary for excitation and that Ca2+ depletion mimics the trp phenotype. We conclude that the recently sequenced trp protein represents a class of light-sensitive channel required for inositide-mediated Ca2+ entry and suggest that this process is necessary for maintained excitation during intense illumination in fly photoreceptors.

Visual transduction in Drosophila.

The brains capacity to analyse and interpret information is limited ultimately by the input it receives. This sets a premium on information capacity of sensory receptors, which can be maximized by optimizing sensitivity, speed and reliability of response. Nowhere is selection pressure for information capacity stronger than in the visual system, where speed and sensitivity can mean the difference between life and death. Phototransduction in flies represents the fastest G-protein-signalling cascade known. Analysis in Drosophila has revealed many of the underlying molecular strategies, leading to the discovery and characterization of signalling molecules of widespread importance.

Polyunsaturated fatty acids activate the Drosophila light-sensitive channels TRP and TRPL.

Phototransduction in invertebrate microvillar photoreceptors is thought to be mediated by the activation of phospholipase C (PLC), but how this leads to gating of the light-sensitive channels is unknown,. Most attention has focused on inositol-1,4,5-trisphosphate, a second messenger produced by PLC from phosphatidylinositol-4,5-bisphosphate; however, PLC also generates diacylglycerol, a potential precursor for several polyunsaturated fatty acids, such as arachidonic acid and linolenic acid. Here we show that both of these fatty acids reversibly activate native light-sensitive channels (transient receptor potential (TRP) and TRP-like (TRPL)) in Drosophila photoreceptors as well as recombinant TRPL channels expressed in Drosophila S2 cells. Recombinant channels are activated rapidly in both whole-cell recordings and inside-out patches, with a half-maximal effector concentration for linolenic acid of ∼10 µM. Four different lipoxygenase inhibitors, which might be expected to lead to build-up of endogenous fatty acids, also activate native TRP and TRPL channels in intact photoreceptors. As arachidonic acid may not be found in Drosophila, we suggest that another polyunsaturated fatty acid, such as linolenic acid, may be a messenger of excitation in Drosophila photoreceptors.

Facets of Vision

This book is a comprehensive review providing an overview of modern compound eye research. The historical development of the field is also sketched, referring in particular to the pioneering study of Sigmund Exner - whose 1891 monograph is at last available in English (please see below). Facets of Vision is also intended as a tribute to Professor H. Autrum, one of this centurys major pioneers of the field. The present state of compound eye research is reviewed by acknowledged international experts, starting with optics and proceeding, through neural processing, to visually guided behaviour. Topics covered include: fundamental optical principles and diversity of optical design, photochemistry, phototransduction, eye pigments, neuroanatomy, circadian rhythms, early visual processing, neuropharmacology, colour vision, polarization vision, depth vision and motion detection. Comprising a coherent compilation of authoritative reviews, Facets of Vision will stand as a milestone in compound eye research.

Common strategies for light adaptation in the peripheral visual systems of fly and dragonfly

Summary1.Intracellular recordings from photoreceptors and large monopolar cells (LMCs) of the flyCalliphora stygia, and the dragonflyHemicordulia tau, were used to examine the peripheral light adaptation processes of the insect compound eye.2.Photoreceptor and lamina adaptation mechanisms were separated by comparing the response waveforms and intensity/response functions (plotted as V/log I curves) of receptors (Figs. 1 and 3) and LMCs (Figs. 2 and 4), subjected to identical regimes of adaptation.3.Photoreceptor adaptation occurs in two phases, a rapid one lasting 100 ms, and a slow phase taking up to 60 s to complete (Fig. 1). This adaptation shifts theV/logI curves to higher intensities without changing their shape or slope (Fig. 3). Adaptation is negligible at low intensities but with stronger adaptation range sensitivity changes approach proportionality to background increments (Fig. 7).4.Lamina adaptation mechanisms adjust the LMCV/logI curve in response to new background levels within 200 ms, producing a phasic response waveform within which background signals are annihilated (Figs. 1, 3, 8). The shape and amplitude of the saturated LMC ‘on’ and ‘off’ transient responses change with light adaptation (Figs. 2, 3).5.At all background intensities examined the slopes of the LMC V/log I curves remain about 8–10 times that of the photoreceptors under the same conditions, implying that lamina adaptation does not change the voltage gain of the first synapse. We propose that light induced depolarisation of the lamina extracellular space subtracts away the standing background signal from the photoreceptor terminals.6.During dark adaptation the faster lamina mechanism can be superimposed upon slower photoreceptor processes (Fig. 9).7.A comparison of our findings with studies of higher order neurons of the compound eye suggests that peripheral adaptation mechanisms play an important role in determining the response of the entire visual system.8.The peripheral light adaptation processes of fly and dragonfly are similar, and the intensity/response functions of retinula cells and LMCs resemble those of vertebrate cones and bipolar cells respectively (Fig. 11). We propose that this analogy has a functional basis. Both vertebrate and invertebrate systems use a ‘log transform-subtraction-multiplication” strategy to match the response bandwidth of peripheral neurons to the expected intensity fluctuation about any one mean, and in so doing maximise the image detail sent to higher centres.

Novel Ca2+ channels underlying transduction in Drosophila photoreceptors: implications for phosphoinositide-mediated Ca2+ mobilization

Drosophila photoreceptors are excellent models for studies of the ubiquitous phosphoinositide signalling cascade. Recent studies suggest that light-induced phosphoinositide hydrolysis in Drosophila leads to the activation of two classes of channels. One is selective for Ca2+ and absent in the transient receptor potential mutant trp. The trp gene product, which shows some structural similarity to vertebrate voltage-gated Ca2+ channels, may thus define a novel family of second-messenger-operated Ca2+ channels generally responsible for the widespread but poorly understood phenomenon of phosphoinositide-mediated Ca2+ entry. The other channel is a non-selective cation channel that requires Ca2+ for activation. As well as being a major charge carrier for the light-induced current, Ca2+ influx via the trp-dependent channels appears to be required for refilling Ca2+ stores sensitive to inositol 1,4,5-trisphosphate and for feedback regulation (light adaptation) of the transduction cascade.

Phototransduction Motifs and Variations

Seeing begins in the photoreceptors, where light is absorbed and signaled to the nervous system. Throughout the animal kingdom, photoreceptors are diverse in design and purpose. Nonetheless, phototransduction-the mechanism by which absorbed photons are converted into an electrical response-is highly conserved and based almost exclusively on a single class of photoproteins, the opsins. In this Review, we survey the G protein-coupled signaling cascades downstream from opsins in photoreceptors across vertebrate and invertebrate species, noting their similarities as well as differences.

Is histamine a neurotransmitter in insect photoreceptors

SummaryIntracellular recordings were made from the large monopolar cells (LMCs) in the first visual neuropil (lamina) of the flyMusca, whilst applying pharmacological agents from a three-barrelled ionophoretic pipette (Fig. 1). Most of the known neurotransmitter candidates (except the neuropeptides) were tested. The LMCs were most sensitive to histamine, saturating with ionophoretic pulses of less than 2 nC. The responses to histamine were fast hyperpolarizations with maximum amplitudes similar to that of the light-induced response (Fig. 3). Like the light response, the histamine response was associated with a conductance increase (Fig. 5). The histamine responses were not blocked by a synaptic blockade induced by ionophoretic application of cobalt ions (Fig. 6). Several histamine antagonists, and also atropine, were effective at blocking or reducing both the response to histamine and the response to light (Fig. 7). Other transmitter candidates having marked effects on the LMCs were: a) the acidic amino-acids, L-aspartate and L-glutamate, which evoked slower hyperpolarizations that could be blocked by cobalt (Fig. 11); b) GABA, which induced a depolarization associated with an inhibition of the light response (Fig. 9); and c) acetylcholine which also caused a depolarization (Fig. 10). Substances with no obvious effect on the LMCs included serotonin (5-HT),β-alanine, dopamine, octopamine, glycine, taurine and noradrenalin. Together with the evidence of Elias and Evans (1983), which shows the presence, synthesis and inactivation of histamine in the retina and optic lobes of the locust, the data suggest that histamine is a neurotransmitter in insect photoreceptors.

Whole-cell recordings of the light induced current in dissociated Drosophila photoreceptors: evidence for feedback by calcium permeating the light-sensitive channels

Tight seal whole-cell recordings of the light-induced current (LIC) were made from photoreceptors in dissociated Drosophila ommatidia. In dark-adapted cells dim light evokes discrete events (quantum bumps) 4‒10 pA in amplitude at resting potential ( ‒ 55 mV). Flash responses scale linearly with light intensity up to an intensity of at least 100 absorbed photons. The dependence of the LIC’s reversal potential on [Ca0] suggests that the channels are primarily permeable to Ca2+ (PCa:PNa ca. 25:1). The current‒voltage relation of the LIC is inwardly rectifying up to reversal potential ( + 10 mV in 0.5 mM Ca0) and then becomes outwardly rectifying. The time to peak increases by up to five times over the range ‒80 to + 60 mV. Raising [Ca0] shortens the time to peak and lessens its dependence on holding potential. It is suggested that there is a sequential positive and negative feedback mediated by Ca2+ ions entering the cell during the light response. In support of this hypothesis, raising [Ca0] during the response first greatly enhances the LIC and then inhibits it. Hyperpolarizing the cell (which will increase the electromotive force for calcium) during the response also results in a transient increase in conductance followed by inhibition. The transient increase in particular can be so rapid (1‒3 ms) that the site of action may be at the channel itself.

Electrophysiological analysis of fly retina. I: Comparative properties of R1-6 and R 7 and 8

Summary1.Intracellular recordings have been made from the photoreceptor classes R1–6, R 7, and R 8 in the flies,Calliphora (wild) andMusca (white).2.The half width of the angular sensitivity function (Δρ) inCalliphora R1–6 varies between 1.5° in the fovea and 3° in the lateral eye regions (Fig. 1). Light adaptation narrows these values by 20%. Both R 7 and R 8 haveΔρ values narrower than in R1–6, averaging 1.3° in foveal regions. InMusca (white)Δρ in R1–6=2.3° and in R7/8=1.5° (both values for foveal regions).3.The presence of an adapting light of sufficient intensity to activate the pupil mechanism shifts the 490 nm peak of spectral sensitivity in R1–6 towards shorter wavelengths by 40–50 nm inCalliphora but not in the pupilless mutant, white, ofMusca (Fig. 2). The shift is independent of photopigment equilibria and has a time course similar to that of the pupil closure mechanism (Fig. 3).4.There are two spectral classes of R7. Both have a single peak of sensitivity in the range 340–360 nm, but one is purely ultra-violet (UV) sensitive, having less than 10% sensitivity beyond 400 nm, whilst the other has a long tail of sensitivity extending to 500 nm. The majority of R 8 cells have a major peak of sensitivity at around 540 nm (Fig. 4).5.Polarisation sensitivity (PS) inCalliphora R1–6 averages 2.0 when measured in the green, and is not significantly affected by light-adaptation. InMusca R1–6, PS averages 1.9. In R7, PS is similar inCalliphora (average 2.2), but reaches values of up to 6 inMusca. PS could not always be detected in R8 but values of up to 3.5 have been recorded inCalliphora.6.Absolute sensitivities measured from the quantal flux required to generate a 50% maximum response using axial light of peak wavelength (APS50) are higher in R 7 and R 8 due to the higher voltage gain per quantum (Table 2).7.All receptor classes light adapt in a similar manner and continue responding to increments of intensity under the brightest adapting regimes used.8.These new results from identified receptors lead to a reappraisal of the possible roles of R1–6 and R 7 and 8 in optomotor responses elicited under different regimes of test pattern wavelength and intensity.