Two-Channel Polarization Analyzer in the Sustaining Fiber-Dimming Fiber Ensemble of Crayfish Visual System

Raymon M. Glantz and Andy McIsaac

Department of Biochemistry and Cell Biology, Rice University, Houston, Texas 77005

    ABSTRACT
Abstract
Introduction
Methods
Results
Discussion
References

Glantz, Raymon M. and Andy McIsaac. Two-channel polarization analyzer in the sustaining fiber-dimming fiber ensemble of crayfish visual system. J. Neurophysiol. 80: 2571-2583, 1998. Polarization sensitivity (PS) was examined in two classes of neurons, sustaining fibers and dimming fibers, in the medulla externa (second optic neuropile) of the crayfish, Pacifasticus leniusculus. Visual responses were recorded intracellularly and extracellularly. The influence of e-vector orientation (theta ) was probed in steady-state responses, with brief flashes and with a rotating polarizer. The results indicate that the entire sustaining fiber population appears to be maximally sensitive to vertically polarized light. Although the evidence is less complete for dimming fibers, they appear to be maximally inhibited by vertically polarized light and excited by horizontally polarized light. Thus the sustaining fibers and dimming fibers form a two-channel polarization analyzer that captures the main features of the polarization system established in photoreceptors and lamina monopolar cells. The available evidence suggests that this two-channel system has the same characteristics across most or all of the retinula. Lateral inhibition in sustaining fibers is differentially sensitive to theta . Inhibition is substantial at theta  = 90° (horizontal) and essentially absent at theta  = 0°. The details of the sustaining fiber polarization response closely follow features established in more peripheral neurons, including the magnitude of PS, enhanced responsiveness to a changing e-vector, and modest directionality to a changing e-vector in~40% of the cells.

    INTRODUCTION
Abstract
Introduction
Methods
Results
Discussion
References

A notable feature of many arthropod and cephalopod visual systems is the sensitivity to the e-vector orientation (theta ) of polarized light (Shashar and Cronin 1996; Waterman 1981, 1984; Wehner 1989). This capacity arises from photoreceptors structurally specialized for the discrimination of theta  (Shaw 1966, 1969; Waterman 1981). Arthropod retinula may contain two to four distinct e-vector channels (Marshall et al. 1991; Shaw 1966; Wehner 1983). Furthermore, polarization analysis may be extended by rotation of the retinula via head and eyestalk movements.

Polarization sensitivity (PS) is expressed in a wide range of behaviors, including navigation (Wehner 1989), the detection of specular reflections (Schwind 1991), and optomotor reflexes (Schöne and Schöne 1961; von Philipsborn and Labhart 1990; Wolf et al. 1980). The neuronal mechanisms that encode polarization-related signals are only partially understood. In crayfish, the retinular cells and lamina monopolar neurons form the basis of two parallel pathways that signal orthogonal theta s (Glantz 1996a; Nässel and Waterman 1977; Sabra and Glantz 1985; Waterman and Fernandez 1970). A subsequent stage consists of "tangential cells," which are components of the lamina-medulla externa (second optic neuropile) circuitry (Strausfeld and Nässel 1981; Wang-Bennett and Glantz 1987b). In one subclass of these cells PS is enhanced by an opponency mechanism, and the second subclass is directionally selective for a changing e-vector (Glantz 1996b). Tangential cells modulate the visual signals of the sustaining fibers (tonic ON neurons) (Wang-Bennett and Glantz 1987b), which in turn synapse on optomotor neurons (Glantz and Nudelman 1988; Glantz et al. 1984). The sustaining fibers and dimming fibers (tonic OFF neurons) are the principal output neurons of the medulla externa (Kirk et al. 1982; Wiersma and Yamaguchi 1966, 1967).

Previous studies of PS in sustaining fibers of shrimp (Yamaguchi et al. 1976) and the crayfish Procambarus clarkii (Yamaguchi et al. in Waterman 1984) indicate strong sensitivity to a changing e-vector. PS was not examined in the dimming fibers. This study describes the stationary and dynamic polarization response of sustaining fibers and dimming fibers in Pacifasticus leniusculus. The results indicate a stationary PS profile with a single peak near the vertical for the entire population of sustaining fibers. Conversely, dimming fibers are inhibited by vertically polarized light and discharge maximally at a horizontal theta .

    METHODS
Abstract
Introduction
Methods
Results
Discussion
References

This study is based on intracellular and extracellular recordings in the crayfish Pacifasticus leniusculus. The animals were prepared as in Glantz (1996b). The eyestalks were glued in their sockets with cyanoacrylate adhesive. The blood was replaced with oxygenated saline during a 1-h perfusion at 8°C. The optic lobe was exposed by excision of the dorsal cuticle of the eyestalk. The animal was clamped in a Plexiglas chamber (containing 2 frosted glass windows, 2 cm on a side) and bathed in oxygenated saline at 20°C. One of the eyes was centered in front of a window at 15 mm from the frosted glass.

Recording procedures

Intracellular recordings from sustaining fibers were made with micropipettes filled with 3.0 M potassium acetate (tip resistances of 100-200 MOmega ) and led to an Axoclamp B1 amplifier. The signals were stored on magnetic tape. All cells were impaled in the medulla externa (Kirk et al. 1982). The same procedures were also used to characterize dimming fibers and medullary amacrine neurons (Waldrop and Glantz 1985).

Extracellular recordings from sustaining fiber axons in the optic nerve were made with tungsten electrodes, sharpened electrolytically to 5.0 µm, and coated with an epoxy-based insulating varnish. The signals were led to an AC amplifier and stored on magnetic tape. For these recordings animals were suspended from a dorsal clamp that could be rotated relative to the optical axis of the stimulus system.


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FIG. 1. Response of sustaining fiber 019 to a changing e-vector orientation (theta ) [clockwise (CW) polarizer rotation] and to stationary theta s. Membrane resting potential is -65 mV and is indicated by zero on the left-hand ordinate. Bottom trace monitors polarizer rotation. theta  = 0° (vertical) is up, and orientation is read from the right-hand ordinate.


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FIG. 2. Mean impulse rates vs. e-vector angle of cell described in Fig. 1. A: responses to stationary e-vector. Rates in pulses/s (pps) are based on 5.0-s samples between 3 and 8 s after each new theta  is established; n = 10. Circles are at ±1.0 SD B: impulse rate vs. time as polarizer is rotated through two 360° cycles (bottom trace). Bottom trace, bullet : e-vector angle of 150°. Scale for e-vector angle is on the right-hand ordinate. C: dynamic phase associated with peak impulse rate vs. stationary e-vector angle of maximum response (theta max) for 21 sustaining fibers. Note 4 data points are superposed. Continuous line is the least-squares regression with slope of 0.95. Correlation coefficient is 0.86.


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FIG. 3. Sustaining fiber (01) polarization response profiles to stationary flashes at varied e-vector angle. A: subthreshold responses. Log I = -4.7. Membrane resting potential is -60 mV and indicated by zero on the left-hand ordinate. Bottom trace: e-vector angle (right-hand ordinate) CW rotation. B: poststimulus time histograms (PSTs) of suprathreshold impulse trains elicited at 15° theta  intervals. theta max is 15°. Each PST is based on 20 responses to 0.5-s flashes, and theta  was varied from -40 to 120°. The peak transient discharge of each PST is aligned with the stimulus theta  on the abscissa. C: PSTs at theta  = 0° for 5 stimulus intensities. PS for the responses in B are determined from the ratio of intensities required for equal magnitude responses at theta max and theta min. D: polarization response profile of another cell (sustaining fiber 014). Note peaks at theta s of 20 and 100°. Data are organized as in B.

Stimulus system

Visual stimuli were generated by two optical channels alternately accessible via a 45° mirror. One channel, consisting of a 7.0-mW helium-neon laser and directed by galvanometer controlled mirrors, was used to locate the receptive field. The sustaining fibers were identified on the basis of receptive field location, after the methods of Wiersma and Yamaguchi (1967). The stimulus was controlled with an electromagnetic shutter and neutral density wedge.

The second channel, used to deliver linearly polarized light, consisted of a 300-W quartz iodide lamp, heat filter, electromagnetic shutter, neutral density wedge, focusing optics, and polarizer mounted on a motorized rotation device. All optical elements were aligned on a single axis. A diaphragm controlled the spot size from 3.0 to 15 mm (11-53°). The focusing lens was mounted on a manipulator to control the target spot location. An optical baffle restricted the illumination of the frosted glass plate to the optic axis and blocked reflected light. The stimulus was monitored with a photodiode covered with vertically oriented polarizer film.

The stimulus consisted of white light, and the unattenuated power was 1.2 mW/cm2, over the wavelength range of 450-650 nm. For most experiments, the diaphragm was set to deliver a 3.0-mm diameter spot, with total maximum power of 0.34 mW.

The optical system was calibrated with a silicon photodiode and polarization analyzer. With the diode placed at the normal position of the crayfish eye, the degree of polarization exceeded 99%, and stray polarization was <0.5%. Stray polarization was measured by rotating the polarizer and measuring the photodiode response with the polarization analyzer removed.

Experimental protocols

Because previous reports (Leggett 1976; Waterman 1984; Yamaguchi et al. 1976) indicated that crustacean visual neurons are insensitive to the e-vector angle of a stationary flash, two protocols were developed to explore this issue. These procedures examine the steady-state and transient responses to variations in e-vector angle.

In the contrast method the eye is continuously exposed to polarized light of constant intensity, and the e-vector angle is varied stepwise. Each theta  is presented for 8-10 s, and the polarizer is then rotated to another angle. Each rotation is associated with a transient burst of~1.0 s in duration. The theta -dependent variations in mean rate were assessed for the interval from 3.0 to 8.0 s after the establishment of each theta . The responses to each theta  were measured 10 times.

Transient responsiveness to theta  was measured with a scanning procedure. Responses were elicited with brief (0.5-0.7 s) pulses of illumination, and the polarizer was rotated to successive angles in steps of 12-22°. For synaptic potential measurements, the responses to four to six stimulus cycles were averaged. For impulse trains, the responses to 20 cycles were averaged to form poststimulus time histograms (PSTs) at each e-vector angle. To avoid saturation, these experiments were performed at intensities <10% of the saturation intensity.

PS was estimated from the response functions by determining the difference in stimulus intensity at theta max necessary to generate comparable responses at theta max and theta min. To determine the dependence of PS on stimulus intensity (I), the response-intensity functions were measured at theta max and theta min. For any response magnitude PS I(theta min)/I(theta max). For each neuron the average magnitude of PS was determined for the intensity range from 0.5 log units above threshold to just below saturation intensity. The PS of dimming fibers was measured in seven cells from the size of the light-elicited inhibitory postsynaptic potential (IPSP).

The response to temporal variations in theta  was examined with a rotating polarizer. The influence of both the rate and direction of change was examined in 30 preparations. The responses were evaluated from the phase and amplitude of the oscillatory membrane potential and from variations in the impulse rate relative to theta .

For studies of lateral inhibition the stimulus system was reconfigured such that two optical pathways, separated by a visual angle of 90°, converged on the eye. Each path consisted of a tungsten lamp, shutter, neutral density wedge, focusing optics, and polarizer. The excitatory stimulus was a slowly rotating polarizer. The stimulus to the inhibitory surround was a constant illumination at fixed theta .

Baseline firing rates were measured with a digital counter that provided indications of drift in excitability or the occurrence of an excited state (Wiersma and Yamaguchi 1967). During excited states measurements were suspended until the control baseline rate resumed.

Data acquisition and analysis

Data acquisition was performed with a Data Translation A/D card and a PC computer. Intracellular recordings were digitized at 500 Hz. For extracellular recordings the action potentials were isolated with a time-amplitude window discriminator and digitized at 300 Hz, which is the maximum discharge rate.

Data analysis was performed with programs written in MATLAB. For intracellular recordings a Butterworth low-pass filter (tau  = 0.03 s) separated action potentials from the compound excitatory postsynaptic potential (EPSP). The filtered signals were then averaged with the stimulus trace as a temporal reference. The action potential sequences were converted into a train of unitary events and used to derive mean impulse rates and to form PSTs. Temporal variations in the impulse rate were derived from the impulse probability divided by the PST binwidth. Extracellular recordings were evaluated from their mean impulse rates and from PSTs.


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FIG. 4. Response-intensity functions of sustaining fiber EPSP at theta  = 0 and 90°. A: signal-averaged EPSP of sustaining fiber 019 for log intensities -4.0 to -1.0 in 0.5 log unit steps at theta  = 0°. Flashes indicated by bar at bottom of figure and were 0.4 s in duration. Each trace is the average of 5 responses. Impulses were removed with a low-pass filter before signal averaging. B: as in A for theta  = 90°. C: transient response amplitude vs. intensity at theta  = 0° () and theta  = 90° (- - -). D: plateau phase amplitude at theta  = 0° () and 90° (- - -). E: mean impulse rate vs. log intensity at theta  = 0° () and 90° (- - -). Vertical bars are ±1.0 SD.

    RESULTS
Abstract
Introduction
Methods
Results
Discussion
References

PS in the response to a stationary e-vector

Rotating a polarizer elicits transient responses, as in Fig. 1 (first 25 s), synchronized to the changes in e-vector angle. As theta  approaches 0°, the membrane depolarizes, and there is a burst of impulses. As theta  approaches 90°, the membrane repolarizes, and the impulse rate declines toward a minimum. Modest changes in e-vector angle (10-30°), produce changes in membrane potential and in mean impulse rate that persist for <= 20 s, as shown in Fig. 1. At each stepwise change in theta  between 0 and 60° the membrane potential and the impulse rate decline. Conversely, successive steps between 60 and 180° produce depolarization and increased impulse rates. Figure 2A shows the mean rate (subsequent to the transient burst) for each of 6 theta s. At theta max (between 150 and 0°) the mean rate was 3.6 times that at theta min (60°), and PS was 7.0.

For purposes of comparison, the variations in impulse rate elicited with a continuously rotating polarizer (at the same intensity) are shown in Figs. 1 (1st 25 s) and 2B. These data exhibit theta max and theta min near 150° (bullet ) and 60°, respectively, and a 3.4-fold difference in responsiveness. Thus continuous rotation augments the discharge, but the ratio of impulse rates at theta max and theta min are similar. The similarity between theta max measured at stationary e-vector angles and the phase of the peak response elicited with a slowly rotating polarizer was observed for nearly all neurons so examined (as in Fig. 2C). Thus, when a crayfish rotates in a field of polarized light, the elements of the sustaining fiber ensemble will each discharge at a rate proportional to the proximity to theta max.

The stationary polarization response profile was also measured with the scanning method, as shown in Fig. 3. In Fig. 3A the data were derived with subthreshold light pulses, and the EPSPs vary from 4.5 mV at theta max (0°) to 2.0 mV at theta min. At suprathreshold intensities the discharge is evaluated with the PST, and PS is interpolated from the intensity-response function. Figure 3B shows theta -dependent variations in firing rate. The peak impulse rate at theta max (15°) is about 2.2 times that at theta min (105°), and PS is 4.0 as determined by the variations in the intensity dependence of the peak rate in Fig. 3C. At moderate intensities, 3 of 33 cells tested exhibited a two-peak sensitivity profile, as in Fig. 3D, with the peaks spaced ~90° apart. An additional five cells exhibited this pattern at higher intensities. Thus there is evidence for convergent input from the orthogonal channels in 10-25% of the cells.

The intensity dependence of PS can be evaluated from intensity-response functions at theta max and theta min, as shown in Fig. 4. In this cell PS increases with stimulus intensity. The responses in Fig. 4, A and B, are averaged compound EPSPs at theta max and theta min. At the lowest intensities, both the transient and plateau responses exhibit only modest PS (<= 2.0, as in Fig. 4, C and D). At higher intensities however the response to flashes at theta  = 0° is saturated yet remains 30-50% greater than the response to flashes at theta  = 90°. This result implies an intensity dependence in PS. The discontinuity at log I = -3.0 may reflect a high-threshold opponency mechanism. Sustaining fibers are subject to intensity-dependent lateral inhibition (Glantz and Nudelman 1976; Waldrop and Glantz 1985). At high stimulus intensities, the EPSP traces in Fig. 4, A and B, exhibit a dip in the membrane potential (arrow in Fig. 4A) between the peak and plateau phases of the response. A similar dip in the PST has been previously shown (Glantz and Nudelman 1976) to reflect a delayed action from the inhibitory surround. Furthermore, the dip occurs over the same range of stimulus intensities associated with the divergence of the response functions at theta max and theta min (in Fig. 4, C and D). More direct evidence for PS in the surround inhibitory mechanism is described below.

The impulse rates between threshold, log I = -3, and saturation (as in Fig. 4E) exhibit an average PS of 5.6 ± 2.0 (SD). Intensity-response functions at theta max and theta min were examined in 12 neurons and revealed an average PS of 4.8 ± 3.0, which is comparable with the PS of receptors and lamina cells in Pacifasticus (Glantz 1996a).

Distribution of theta max in sustaining and dimming fibers

The theta max of sustaining fibers clusters near the vertical as previously inferred by Yamaguchi et al. (1976) from dynamic responses. The distribution of theta max values for 48 cells measured with a stationary polarizer is shown in Fig. 5A. The data sample includes 9 of 14 sustaining fiber subtypes (receptive field locations) identified by Wiersma and Yamaguchi (1967). When these results are segregated by receptive field location (dorsal vs. ventral, anterior vs. posterior) the result is similar for each subgroup and similar to Fig. 5A (data not shown). A striking feature of the distribution is the paucity of theta max observations near 90°. Only 2 of 48 cells exhibited theta max within ±15° of the horizontal. This distribution contrasts markedly with those of receptors and lamina monopolar cells (Glantz 1996a) in which both orthogonal theta max values are well represented in the cell populations. The preference for a vertical e-vector is quite robust. When the stimulus is centered in the inhibitory surround such that only the internally reflected light reaches the excitatory field, the distribution of e-vector maxima remains clustered about the vertical. An interesting feature of PS in dimming fibers is that the distribution of theta max for inhibition, Fig. 5B, is similar to that for excitation in sustaining fibers.


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FIG. 5. A: distribution of theta max values from stationary flash measurements for 48 sustaining fibers. B: distribution of theta max values for the elicited inhibitory postsynaptic potential (IPSP) of 14 dimming fibers. Seven measurements were based on the scanning method, and 7 were based on the theta  associated with maximum hyperpolarization in response to a slowing rotating polarizer.

Sustaining fiber response to a changing e-vector

The sustaining fiber response to temporal variations in theta  reflects both the stationary PS profile and the direction and rate of change of theta  (as in Figs. 1 and 2). Figure 6 shows the subthreshold response to polarizer rotation at 76 and 146°/s in the clockwise (CW) and counterclockwise (CC) directions. The oscillations in synaptic potential are modestly larger (30-50%) for CC rotations and at the higher velocity. Directionality and sensitivity to rates of change were examined in 30 neurons. Thirteen cells exhibited a consistent directional bias similar to that shown in Fig. 6 and with responses in one direction exceeding that in the opposite direction by 40-100%. Most cells were also sensitive to the rate of change of the e-vector, with a maximum response at 100-180°/s and diminished responses at lower and higher rates of change (data not shown).


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FIG. 6. Sustaining fiber 01 subthreshold response to a rotating polarizer. The shutter is opened at t = 2.2 s (bottom trace) eliciting a transient depolarization that decays to a plateau phase (top trace). The membrane resting potential was -60 mV, and it is indicated by zero on the left-hand ordinate. Polarizer rotation commences at t = 12 s in the CW direction at 76°/s, followed by counterclockwise (CC) rotation at 76 and 146°/s and CW rotation at 146°/s. Bottom trace is stimulus monitor, and e-vector angle is indicated on the right-hand ordinate.

PS in the inhibitory surround

The intensity dependence of PS described in Fig. 4 and the insensitivity of theta max to stimulus location may have a common basis in sustaining fiber surround inhibition (Waldrop and Glantz 1985; Wiersma and Yamaguchi 1967). If the inhibitory mechanism is polarization sensitive with theta max near 90°, the PS profiles for net excitation would be shifted toward 0°. The results of two types of experiments suggest that the sustaining fiber inhibitory mechanism is polarization sensitive and that theta max for inhibition is ~90°.

In one procedure, theta max for excitation was assessed with a rotating polarizer (30-60°/s) and with the light beam focused on the excitatory field. After a control measurement (as in Fig. 7A) the inhibitory field was illuminated with constant polarized light from a second beam at theta  = 0° or theta  = 90°, as seen in Fig. 7, B and C, respectively. When the inhibitory stimulus is polarized to theta  = 0°, there is little or no evidence of inhibition. When the inhibitory field was illuminated at theta  = 90°, both the peak and mean rates, elicited by the excitatory stimulus, were substantially diminished. Similar results were obtained in three of four experiments. An inhibitory stimulus at theta  = 90° was 20-50% more effective than one at 0°. The fourth cell revealed no evidence of inhibition at any theta .


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FIG. 7. Dependence of lateral inhibition on theta . A: control response of sustaining fiber 038 (left-hand ordinate) to CW polarizer rotation (bottom trace, right-hand ordinate). Mean rate is 10.9 ± 0.1 impulse/s. B: as in A but with continuous illumination of the inhibitory surround at theta  = 0°. Mean rate is 9.7 ± 1.1 impulses/s. C: as in B but illumination of inhibitory field is at theta  = 90°. Mean rate is 5.3 ± 2.3 impulses/s.

In a second experiment, the PS of the cells that mediate sustaining fiber lateral inhibition (Fig. 12), the medullary amacrine cells (Waldrop and Glantz 1985), was examined with a stationary or slowly rotating polarizer as in Fig. 8. In this cell, theta max was at 78°, as shown in Fig. 8A. Three of six cells exhibited a theta max near 90°, and only one revealed a theta max near 0°. The theta max, evaluated from the response to a rotating polarizer, depends on the e-vector rate of change. A theta max near 90° obtains at rates of up to 120°/s (as in Fig. 8B) but not at higher rates (as in Fig. 8, C and D).


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FIG. 12. Schematic diagram of connections in the crayfish visual system related to PS. RH and RV are retinular cells sensitive to the horizontal and vertical e-vector angles, respectively. There are 4 RVs and 3 RHs in each ommatidium. Lamina monopolar cells M3 [1 per lamina (LM) cartridge] receive sign-inverting, receptor input exclusively from 4 RVs originating in a single ommatidium (Nässel and Waterman 1977), and M4 is exclusively innervated by 3 RH cells per ommatidium. Monopolar cells project retinotopically to the medulla externa (ME) and synapse on columnar transmedullary neurons (TM). Cholinergic transmedullary cells (TM-A) with horizontal e-vector sensitivity (innervated by M4) provide depolarizing (sign-conserved) synaptic input to broad field medullary amacrine neurons. TM-A with a vertical e-vector preference (innervated by M3) provides a sign-inverting (hyperpolarizing) input to dimming fibers. Sustaining fibers are excited by glutamatergic transmedullary cells (TM-GL) with V-sensitivity. TM-GL are inhibited by medullary amacrine cells, which is the basis of the H-sensitivity in sustaining fiber lateral inhibition. In the brain [supraesophageal ganglion (SG)], sustaining fibers excite optomotor neurons or indirectly inhibit motoneurons via nonspiking interneurons.


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FIG. 8. Response of nonspiking medullary amacrine neuron to CW polarizer rotation at 53°/s (A), 120°/s (B), 218°/s (C), and 300°/s (D). Note at the lowest velocity the maximum depolarization obtains at theta  = 78°, and phase lag increases at successively higher rates of e-vector change. Each trace is the average of 5 (A) or 10 (B-D) responses. Membrane resting potential is -50 mV, indicated by 0 on left-hand ordinate. Broken traces are from the stimulus monitor and scaled on the right-hand ordinate.

PS in dimming fibers

Dimming fibers are spontaneously active in the dark and exhibit an OFF response to intensity decrements. Increments of illumination elicit a graded hyperpolarization and a pause in the discharge, as shown in Fig. 9. With polarized light, the hyperpolarizations elicited by pulses at theta  = 0° (as in Fig. 9, right column) are larger than those obtained with pulses at 90° (as in Fig. 9, left column), and the theta -dependent difference generally increases with stimulus intensity, as in Fig. 9G.


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FIG. 9. Dimming fiber IPSPs elicited with 2.0-s pulses of light at theta  = 90° (left column) and 0° (right column) and at log intensity -3 (A and B), -2 (C and D), and -1 (E and F). Note that the IPSP increases progressively with intensity at theta  = 0° but not at theta  = 90°. Lower bar indicates stimulus timing. G: IPSP amplitude vs. intensity at theta  = 0° () and theta  = 90° (- - -). Circles are at ±1.0 SD.

The dimming fiber polarization response profile (as in Fig. 10A), is a continuous function of theta  and theta max for inhibition is ~0°. The IPSP at theta  of 0° is ~60% larger than that at 90°, and the response varies as a cos2 theta  function, as in Fig. 10B. The average PS of dimming fibers was 4.9 ± 3.0 (n = 7). theta max for inhibition was determined with the scanning method, as in Fig. 10, or from the response to a slowly rotating polarizer, as in Fig. 11. In 12 of 14 cells so tested, theta max was within ±15° of 0°, as in Fig. 5B. Thus dimming fibers are inhibited, and sustaining fibers are excited at the same theta max. For a changing e-vector, inhibition at theta  = 0° hyperpolarizes the cell as theta  approaches 0°, and the cell depolarizes (recovers from inhibition) as theta  approaches 90°, as shown in Fig. 11. Although the theta -dependent variation in membrane potential is a modest 1.0-3.0 mV (as in Fig. 11, B and C), it is sufficient to maximize the impulse rate as theta  approaches 90°.


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FIG. 10. A: dimming fiber responses to flashes (0.6-s duration, at 2-s intervals) of constant intensity and with theta  varied in 17 steps per 180°. Log I = -1.0. Lower trace is the stimulus monitor, and stimulus scale is on the right-hand ordinate. Polarizer rotation is CW. B: average polarization response profile for the dimming fiber IPSP at log I = -1.0. : mean response amplitude for 5 stimulus cycles. open circle  are at ±SE. - - -: cos2 theta  function with theta max of 11°. PS for this neuron was 2.4, and comparable response ratios were obtained at log I = -1.5 and -2.0.


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FIG. 11. A: dimming fiber response to polarizer rotation at 45°/s. Note depolarization and peak impulse rate near theta  = 90°. Membrane resting potential is -54 mV, indicated by 0 on left-hand ordinate. Dashed line monitors the e-vector angle (right-hand ordinate). Rotation is CW. B and C: signal averages of synaptic potential responses to polarizer rotation at 30 and 67°/s, respectively. Note maximum hyperpolarization near theta  = 0°.

    DISCUSSION
Abstract
Introduction
Methods
Results
Discussion
References

The principal purpose of this study was to examine the transfer of polarization-related information from the more distal nonspiking visual neurons to the sustaining fibers and dimming fibers. Toward this end, aspects of PS, previously measured in lamina monopolar cells and tangential cells, were examined with the same procedures in sustaining and dimming fibers. Both monopolar cells and tangential cells make indirect functional connections to sustaining fibers (Wang-Bennett and Glantz 1987a,b).

The monopolar cells exhibit an average PS of 4.5 (Table 1); they provide separate pathways for horizontal and vertical theta s (Fig. 12) and enhanced responsiveness to a changing e-vector. The sustaining fiber polarization response is similar to that of only one class of lamina monopolars (with theta max ~0°, as in Table 1). The absence of a horizontal e-vector channel in sustaining fibers implies a highly selective pattern of innervation. Because the dimming fiber theta max at 90° results from inhibition at theta  = 0°, the dimming fiber input must also come from a subset of columnar cells with theta max = 0°. Excitation of sustaining fibers is mediated by glutamate (Pfeiffer-Linn and Glantz 1991), but inhibition of dimming fibers requires acetylcholine (Pfeiffer and Glantz 1989). Thus the two synaptic actions, which have similar theta  dependence, must be mediated by different transmedullary cells, as shown in Fig. 12. It is likely however, that these cholinergic and glutamatergic transmedullary cells are excited by common lamina monopolar neurons (M3 in Fig. 12). Another inference from these studies is that the glutamatergic transmedullary cells (Pfeiffer-Linn and Glantz 1991), which are presynaptic to sustaining fibers (TM-GL, in Fig. 12), are likely to exhibit a theta max near 0°.

 
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TABLE 1. Comparison of the polarization-sensitive properties in four classes of neurons

Sustaining fiber PS also resembles that in one class of tangential neuron (type I), which exhibits modest directionality in the response to a changing e-vector. Thus sustaining fibers appear to capture the essential features of a subset of lamina monopolar neurons and the type I tangential cells.

The results indicate that the output side of the medulla externa contains two pathways with orthogonal e-vector sensitivity. The signals are transmitted to the brain via their axons in the optic nerve (as in Fig. 12). The essential features of the photoreceptor-lamina monopolar PS system are all contained in the sustaining-dimming fiber ensemble. These include the stationary PS profiles of two orthogonal channels, enhanced responsiveness to a changing e-vector, the fidelity of dynamic phase and stationary theta max, and the representation of PS for the entire panoramic visual field.

The function of the PS system for crayfish behavior is less clear. One problem is that all the neurons of the polarization-sensitive afferent pathway are also sensitive to the local contrast of unpolarized light. Thus a change of theta  in a local area will effect particular sustaining and dimming fibers in a manner that is indistinguishable from a change in local intensity. There are several possible resolutions to this problem, and each requires assumptions about the behavioral context. One possibility is that, in a restricted environment, local intensity and theta  may be correlated. Alternatively, the PS in this system may enhance contrast detection (caused by reflected polarized light) in circumstances in which intensity contrast is absent (Bernard and Wehner 1977; Leggett 1976). In this context it may be immaterial whether the source of the contrast is a difference in theta  or local intensity.

Previous studies described PS in neurons of the crab medulla externa (Leggett 1976). The cells are insensitive to the stationary e-vector angle but highly sensitive to e-vector change and/or the direction of change. In contrast to the crayfish sustaining and dimming fibers, however, the crab neuron's response is not modulated with variations in theta . The opposite extreme is found in the cricket medulla, which contains three classes of polarization-sensitive neurons with theta max separated by ~60° (Labhardt 1988). Stationary PS is enhanced by an opponency mechanism (Labhardt and Petzold 1993). It is notable that three arthropods analyze polarization signals in homologous structures (the second optic neuropile) but with very different strategies.

An important difference regarding PS in crayfish sustaining and dimming fibers and that in several insect species is the distribution of polarization analyzers across the retinula. In crayfish PS is broadly distributed across the retinula, and there are orthogonal e-vector analyzers at all or most locations. In bees, ants, and crickets (Wehner 1989) PS is largely restricted to the ommatidia on the dorsal face of the compound eye, and the distribution of analyzers is consistent with a matched filter for skylight polarization. This organization provides a basis for navigation by polarized skylight. The more homogeneous distribution of PS in crayfish eye may indicate a different function such as contrast enhancement in an aquatic environment.

    ACKNOWLEDGEMENTS

  We thank S. Rabinowitz for assistance in preparation of the manuscript.

  This research was supported by a National Science Foundation Grant IBN-9507878.

    FOOTNOTES

  Address for reprint requests: R. M. Glantz, Dept. of Biochemistry and Cell Biology, Rice University, 6100 Main St., Houston, TX 77005.

  Received 25 August 1997; accepted in final form 23 July 1998.

    REFERENCES
Abstract
Introduction
Methods
Results
Discussion
References

0022-3077/98 $5.00 Copyright ©1998 The American Physiological Society