DOI: 10.5061/dryad.tb2rbnzws
title: The integration of sensory feedback in the modulation of anuran landing preparation

description: Controlled landing requires preparation. Mammals and bipedal birds vary
 how they prepare for landing by predicting the timing and magnitude of
 impact from the integration of visual and non-visual information. Here we
 explore how an animal that moves primarily through hopping, Rhinella
 marina, the cane toad, integrates sensory information to modulate landing
 preparation. Earlier work suggests toads may modulate landing preparation
 using predictions of impact timing and/or magnitude based on non-visual
 sensory feedback during takeoff rather than visual cues about the landing
 itself. Here we disentangled takeoff and landing conditions by hopping
 toads off platforms of different heights and measured electromyographic
 (EMG) activity of an elbow extensor, m. anconeus, and used high-speed
 motion capture to quantify whole body and forelimb kinematics to test how
 toads integrate visual and non-visual information in landing preparation.
 We asked two questions: 1) when they conflict, do toads correlate landing
 preparation with takeoff or landing conditions?  And 2) for hops
 with the same takeoff conditions, does visual information alter the timing
 of landing preparation?  We found that takeoff conditions are a
 better predictor of the onset of landing preparation than landing
 conditions, but that visual information is not ignored. When hopping off
 higher platforms, toads start to prepare for landing later when takeoff
 conditions are invariant. This suggests that, unlike mammals, toads
 prioritize non-visual sensory feedback about takeoff conditions to
 coordinate landing, but that they do integrate visual information to
 fine-tune landing preparation.
description: Animals Six female cane
 toads, <i>R. marina</i>, ranging in mass from 63 to 170 g were
 used for kinematic analysis and electromyography (EMG) recordings. All
 animals were housed in groups of three to four in large aquaria on a 12
 hr. light dark cycle and fed crickets several times weekly.  All
 experimental work was approved by Mount Holyoke College's
 IACUC. Electromyography and kinematic data
 collection EMG data were
 collected from the <i>m. anconeus</i>, an elbow extensor known
 to show distance-dependent pre-landing onset timing (Gillis et al., 2010).
 To implant electrodes, toads were anesthetized in a solution of MS-222
 (1.5 g l-1). Once anesthetized, 1-2 cm skin incisions were made along each
 humerus to expose the muscle. Bilateral implants were used to increase the
 likelihood of successful data collection. Bipolar electrodes were made and
 implanted as described in detail in previous work (Gillis et al., 2010).
 EMG signals were amplified 1000X with Grass P511 preamplifiers using a
 notch filter at 60 Hz. Signals were digitized at 5000 Hz using a NIDAQ
 16-bit A/D converter and a custom LABVIEW program and saved onto a
 personal computer. Toads alter both the timing of
 pre-landing EMG activity and the beginning of elbow extension with hop
 distance.  For completeness, we collected both EMG data and forelimb
 kinematics. For kinematic data collection, small squares of white
 cardboard (~3X3 mm) were glued to the skin bilaterally at the wrist and
 elbow joints as well as at mid-point of the humerus (markers at the
 shoulder joint are readily obscured). In addition, four markers were also
 used to form a T along the back of the animal as described in detail in
 previous work (Cox and Gillis, 2015). Once recovered from anesthesia,
 animals were placed in a well-lit enclosure (64X107 cm) and, in a random
 order, hopped off three different height platforms (0 cm (41 hops), 6 cm
 (36 hops) and 10 cm (31 hops)) lined with felt to ensure purchase. Between
 3 and 12 hops per condition per toad were included in the analysis
 (mean:6.8, SD:3.8 hops/treatment/toad) for a total of 102 hops. Videos of
 hops were taken with two Fastec HiSpec1 high-speed video cameras at 500
 fps. For three-dimensional spatial calibration, a 64-point calibration
 cube (21X21 cm) was digitized in MATLAB using DLTdv5 (Hedrick, 2008).
 Videos were synchronized with EMG signals using a 5V trigger pulse that
 stopped video and EMG recording and was included on its own channel with
 EMG data. After hopping trials, toads were euthanized by overnight
 submersion in MS-222 (1.5 gl-1). Post-mortem dissections were used to
 confirm electrode placements. Data analysis Landing
 preparation in toads is characterized by both distance-dependent amounts
 of elbow extension and activation of the underlying forelimb musculature
 at touchdown. But landing preparation at touchdown is not sufficient to
 determine whether the motor control strategy is altered. Modulation of
 pre-landing EMG activity and elbow configuration at touchdown can be
 achieved without varying the timing or intensity of muscle activation. A
 toad could start elbow extension and muscle activation at the same time
 relative to liftoff and maintain the same rate of elbow extension and
 increase of activation intensity throughout the hop. With this strategy,
 long hops provide more time to generate greater levels of activation or
 elbow extension without modulating the motor control strategy. Thus, to
 discriminate changes in motor control, we chose to primarily focus our
 analysis on how toads changed the timing of landing preparation (the onset
 and duration of pre-landing EMG activity or elbow extension) since the
 timing of prelanding EMG activity for the muscle instrumented varied more
 consistently with hop duration than amplitude. For completeness, we also
 evaluated metrics of the intensity of both pre-landing EMG activity and
 the rate of elbow extension 60 ms after liftoff.  To quantify how
 variations in motor control influence landing preparation at touchdown, we
 also measured EMG intensity over a 60 ms window before landing as well as
 the elbow’s configuration (degree of extension) at touchdown.
 Video
 Analysis: Videos were analyzed to identify the timing of the
 onset of movement (T0), time of takeoff (takeoffs) and landing for each
 limb (ltouchs: touchdown of left limb, rtouchs: touchdown of right limb,
 touchs: time of first limb to touchdown). The onset of movement was
 defined as the moment the toad's vertical velocity first topped 5
 cm/s. Three-dimensional coordinates of the forelimbs were calculated with
 Matlab software (Hedrick, 2008). Data were smoothed with a quintic spline
 interpolation, and elbow angle, the timing of extension onset (E2) and
 extension velocity (EEvel60), as well as the trajectory and
 velocity of the animal during the hop, were calculated as described
 elsewhere from the vertical velocity of the animal at takeoff
 (maxT0velVBB), the takeoff height of the snout of the animal
 (maxT0velHeight) and the height of the animal at reast
 (startHeight). Predictions of touchdown times: The time touchdown would have occurred if animals were hopping on flat ground was calculated with equations of motion from the vertical velocity of the center of mass and height of the marker on the tip of the animal’s snout at takeoff. Time of touchdown, t<sub>TD</sub> was the sum of the time of liftoff, t<sub>LO</sub>, the time to rise to peak height, t<sub>R</sub>, and the time to fall back to starting height, t<sub>F</sub>. <i>t</i><sub><i>TD</i></sub><i>= </i><i>t</i><sub><i>LO</i></sub><i>+</i><i>t</i><sub><i>R</i></sub><i>+</i><i>t</i><sub><i>F</i></sub><i>.</i> Starting height was the height of the tip of snout before hop initiation. The time to peak height, t<sub>R</sub>, was found from the vertical component of the velocity of the center of mass at liftoff, <i>Vz<sub>LO</sub></i>, and acceleration due to gravity.  Given that the vertical velocity at peak height is zero, <i>t</i><sub><i>R </i></sub><i>= </i><i>Vz</i><sub><i>LO</i></sub><i>/g.</i> The time to fall back to start height, t<sub>F</sub>, was calculated from the total height to fall from peak height, h<sub>P</sub>, back to starting height, h<sub>S</sub>. Given that rise height can be found from <i> </i><i>1</i><i>2</i><i>a</i><i>t</i><sup><i>2</i></sup> , the peak height was the sum of the takeoff height, h<sub>LO</sub>, <i>h</i><sub><i>P</i></sub><i>= </i><i>h</i><sub><i>LO</i></sub><i>+ </i><i>g</i><i>t</i><sub><i>R</i></sub><sup><i>2</i></sup><i>2</i><i>.</i> The time to fall, then, is <i>t</i><sub><i>F </i></sub><i>= </i><i>2</i><i>h</i><sub><i>p</i></sub><i>g</i><i>.</i> While several approaches to calculating time of flight were tested, including limb angle at touchdown and equations to estimate anuran jumps (Marsh, 1994), this approach best fit our data for flat hops (Fig. 2B). We evaluated how well our predictions lined up with impact time by comparing predicted to actual impact times for flat hops (Flat hops from Group 1) by fitting two mixed models, a full model with predicted touchdown time as the response variable and actual touchdown time as the fixed effect and a null model with no fixed effect. In both models, individuals were a random factor. The R<sup>2</sup> value for the full model was calculated from the relative likelihood of each model determined by their AIC values (Nakagawa). Electromyography and kinematics: EMG activity was analyzed using customized MATLAB scripts in which the onset timing of pre-landing muscle activity (onanc) was identified visually for each hop and the rectified, integrated area, or intensity, of the EMG signals for the first 60 ms after onset (int60Onset)  and the last 60 ms before landing (intAL60touch) was calculated. To control for differences in electrode construction and placement between individuals, each muscle's EMG intensities were normalized to the largest intensity value observed for each animal.  The onset of elbow extension was determined using methods described elsewhere and elbow extension was calculated at touchdown (TDext). The rate of elbow extension (EE<sub>Vel60</sub>) was the average rate of change of elbow extension during the 60 ms after the onset of elbow extension. Data was filtered to only include signals from the limb that touched down first.
description: ReadMe Notes for
 ‘DataFileCoxGillis2019’. This data
 accompanies the paper “The integration of sensory feedback in the
 modulation of anuran landing preparation”  By SM Cox and GB Gillis
 published in Journal of Experimental Biology 2019   The
 file is a csv file with the following columns: toad                      : animal
 identifier hop                       : hop
 number f: hop on flat ground, l: hop  off 6 cm platform, h: hop of 10 cm
 platform takeoffs               : time
 vertical velocity reaches 5 cm/s (s) ltouchs                 : time
 left forelimb touchdown (s) rtouchs                : time
 right forelimb touched down (s) mass                     : animal
 mass (mg) T0                          :
 start of hop (s) E2t                        : time
 of start of elbow extension in preparation for landing onLanc                 : time of
 start of prelanding EMG activity left forelimb onRanc                : time of
 start of prelanding EMG activity left forelimb dist                        : hop
 distance (cm) height                   :
 vertical height difference from resting to touchdown (cm) TDext                   : elbow
 extension of forelimbs at touchdown (180 = straight limb) treat                     :
 treatment: f: hop on flat ground, l: hop  off 6 cm platform, h: hop of 10
 cm platform touchs                  : time
 first limb touchdown intAL60touch      : integrated EMG
 60 ms before touchdown maxTOvelAngleBB            : angle
 of torso from horizontal at maximum vertical velocity (deg)
 maxTOvelVBB    : maximum vertical
 velocity of snout at takeoff (cm/s) maxTOvelHeight: height of snout of
 the animal at time of maximum vertical velocity during takeoff
 (cm) maxTOvelBBT     : time of maximum
 vertical velocity during takeoff (s) dropHeight          : difference
 between maximum vertical position during hop and landing height
 (cm) startHeight          : height of
 the animal at rest from reference zero LR                          :
 which limb touched down first L: left, R: right onanc                   : timing
 of onset of EMG for limb that touched down first predictTD            : timing of
 predicted touchdown TD60Vel               : elbow
 extension velocity 60 ms before touchdown int60Onset          :Integrated
 EMG activity 60 ms after beginning of elbow extension EEvel60                : Average
 elbow extension 60 ms before touchdown (deg/msec)