Physiological evidence of sensory integration in the electrosensory lateral line lobe of Gnathonemus petersii
Physiological evidence of sensory integration in the electrosensory lateral line lobe of Gnathonemus petersii
Sylvia Fechner 0 1 2
Kirsty Grant 0 2
Gerhard von der Emde 0 1 2
Jacob Engelmann 0 1 2
0 Current address: Stanford School of Medicine, Department of Molecular and Cellular Physiology , Stanford, California , United States of America
1 University of Bonn, Institute for Zoology , Bonn, Germany, 2 UNIC, CNRS, 1 Avenue de la Terrasse, Gif-sur Yvette , France , 3 University of Bielefeld, Biology ± AG Active Sensing , Bielefeld , Germany
2 Editor: Maurice J. Chacron, McGill University Department of Physiology , CANADA
Mormyrid fish rely on reafferent input for active electrolocation. Their electrosensory input consists of phase and amplitude information. These are encoded by differently tuned receptor cells within the Mormyromasts, A- and B-cells, respectively, which are distributed over the animal's body. These convey their information to two topographically ordered medullary zones in the electrosensory lateral line lobe (ELL). The so-called medial zone receives only amplitude information, while the dorsolateral zone receives amplitude and phase information. Using both sources of information, Mormyrid fish can disambiguate electrical impedances. Where and how this disambiguation takes place is presently unclear. We here investigate phase-sensitivity downstream from the electroreceptors. We provide first evidence of phase-sensitivity in the medial zone of ELL. In this zone I-cells consistently decreased their rate to positive phase-shifts (6 of 20 cells) and increased their rate to negative shifts (11/20), while E-cells of the medial zone (3/9) responded oppositely to I-cells. In the dorsolateral zone the responses of E- and I-cells were opposite to those found in the medial zone. Tracer injections revealed interzonal projections that interconnect the dorsolateral and medial zones in a somatotopic manner. In summary, we show that phase information is processed differently in the dorsolateral and the medial zones. This is the first evidence for a mechanism that enhances the contrast between two parallel sensory channels in Mormyrid fish. This could be beneficial for impedance discrimination that ultimately must rely on a subtractive merging of these two sensory streams.
Data Availability Statement: All relevant data are
within the paper and its Supporting Information
Funding: This work was supported by Grant EN
826/4-1 from the Deutsche
Forschungsgemeinschaft, http://www.dfg.de/; and
a travel grant by the Deutscher Akademischer
Austauschdienst (DAAD), https://www.daad.de/de/.
The funders had no role in study design, data
collection and analysis, decision to publish, or
preparation of the manuscript.
Neuronal maps in sensory physiology have been studied from at least two perspectives,
defining a map as a neuronal representation that is based on the topography of the receptor array
and/or as a topographic neuronal representation of features that are computed independently
from the topography of the receptor array. The latter are considered as evidence that neuronal
maps can be beneficial beyond the idea of optimal wiring or developmental constraints. To
which degree neuronal maps are of functional relevance is still unanswered [1±3]. It is
commonly accepted that topographic representations facilitate localisation of spatially sparse
], but no optimal representation of multidimensional inputs in the low-dimensional
space of neurons has been defined yet [5±8].
Parallel (mapped) processing of sensory features is common at early processing stages,
particularly at the level of primary sensory input. This has been studied extensively in the
electrosensory lateral line lobe (ELL) of weakly-electric fishes. These fish can actively generate an
electric field through the discharge of their electric organ (EOD). The environmental
modulations of this field are encoded through an array of cutaneous electroreceptors. This input is
used for active electrolocation [
]. Research on the ELL of Gymnotiform fish has advanced our
understanding of the cellular mechanisms that aid in extracting different features from a single
sensory stream through parallel processing [
]. Here, three parallel topographic maps exist in
which subpopulations of neurons with differing spatiotemporal tuning properties process the
information of the sensory input from a single class of electroreceptors in parallel. The three
ELL maps likely evolved through duplication of a plesiomorphic ampullary or
mechanosensory lateral line map [
] and have been interpreted as adaptations to the increased
behavioural repertoire that electroreception offered these fishes [
]. The alternative option to add
the new computational loads to the existing neuronal architecture apparently led to significant
constraints, thus favouring a duplication of maps [13±16]. Interestingly, the three maps in the
Gymnotiform ELL lack interconnections [
]. At the midbrain their input converges on
multiple ill-defined maps [
] but it is unclear if the input of the maps converges on the single cell
]. Recently it was shown that neurones in the midbrain can extract specific features of
the sensory input that are not being responded to at the earlier levels of the sensory pathway.
This gives support to the notion that convergence of parallel sensory streams can enable the
extraction of specific sensory cues [17±19].
The second family of weakly electric fish, the Mormyridae, allow the investigation of
parallel processing of features that are already separated at the receptor level. This offers the
potential to unravel how merging of parallel sensory streams can aid in the extraction of
behaviourally relevant computed sensory features. Mormyrid electroreceptors (Mormyromasts)
are sensitive to amplitude and waveform modulations of the electric field [
]. Contrary to
Gymnotiformes, two differently tuned sensory cells in each mormyromast, A- and B-cells,
respectively, are responsible for encoding these features [
]. Afferents of A- and B-cells
respond to an increase in the amplitude of the EOD with a decrease of their first-spike latency
and an increase in spike number [
]. B-cell afferents in addition are responsive to the
waveform distortions caused by capacitive objects [
]. Amplitude and waveform
modulations (phase) thus can be considered as two parallel streams of sensory information. As
capacitive and resistive properties of an object can modify the responses in the B-cells, whereas the
A-cells are tuned to the resistive properties only, a direct separation of resistive and capacitive
properties is impossible. This requires a central (subtractive) comparison of both sensory
]. A series of behavioural studies showed that Gnathonemus can indeed
discriminate between resistive and capacitive properties unequivocally [
] and this has further
strengthened the hypothesis that the parallel sensory streams of A- and B-cell input need to be
merged centrally [
A- and B-cell information is processed in the somatotopically organized medial zone (MZ)
and the dorsolateral zone (DLZ) [
] of the ELL. Interzonal connections connect the two
zones homotopically, preserving the topography between zones [
]. Such connections could
aid in the disambiguation of electric phase and amplitude, rendering neurones of the DLZ
sensitive to phase only. However, experiments directly addressing this hypothesis found no
2 / 17
evidence that neurones in the DLZ are sensitive to phase only, nor support for an acquired
waveform-sensitivity in the MZ [
]. Stirred by a recent study that showed that B-cell
information is overrepresented in the dorsolateral map of the ELL for the head and chin appendix
], we decided to re-investigate this. Given that the initial study by von der Emde
and Bell (1994) considered neurones receiving input from the trunk region only, we speculated
that a separation of phase and amplitude might be restricted to the head and chin appendix
region. Hence extracellular single-cell recordings and tracer injections were carried out in the
rostral parts of the DLZ and MZ, while stimulating neurones with artificially modified EODs
that only differed in either waveform or amplitude. We found that phase shifted EODs
influenced the neuronal responses in both zones, strongly suggesting that phase information must
be conveyed to the medial zone. We further show that responses of phase-sensitive neurones
in the DLZ and in the MZ differed in a consistent and zone-specific manner. This differential
responsiveness, most likely mediated by the interzonal connections, results in contrast
enhancement that would be beneficial for the proposed subtractive mechanism required to
discriminate amplitude and phase information.
Material and methods
A total of 17 individuals of the species G. petersii were used in the experiments (8.0±12.5 cm in
standard length). The fish were acquired from a local supplier (Aquarium Glaser, Frankfurt/
Main, Germany) and were kept in groups in 250 l aquaria at 25±27 ÊC on daily 12/12 h light/
dark cycle (water conductivity 90±120 μS cm-1).
Surgery was conducted as previously described [
]. Briefly, fish were anaesthetised with
0.1 g l-1 tricaine (MS-222, Acros organics) followed by an intramuscular injection of 0.3±0.5 μl
g-1 body weight pancuronium bromide (Roth). The fish was placed on a Styrofoam platform
and respirated artificially (MS-222: 0.03 g l-1). Before removing dorsal parts of the skin and
skull to expose the valvula cerebelli, the skin was locally anaesthetised (Xylocaine gel 2%,
AstraZeneca, Wedel, Germany). After surgery, the respiration was switched to fresh aerated
water and the Styrofoam platform was removed so that the fish was only held by a plastic rod
attached to the skull.
The study was carried out in accordance with the recommendations in the Guide for the Care
and Use of Laboratory Animals of the National Institutes of Health, with the council of Europe
Treaty ETS 123 as well as with the current local laws of Germany where the experiments were
performed. The protocol was approved by the Landesamt fuÈr Natur, Umwelt und
Verbraucherschutz Nordrhein-Westfalen (LANUV, permit 50.203.2-BN7/107). All surgery was
performed under sodium bicarbonate buffered tricaine methanesulfonate anaesthesia; in addition
local analgesia was administered at the wound margins (lidocaine), and all efforts were made
to minimize suffering. Monitoring of animal welfare during the experiments was conducted
]). When fish ceased to produce EODs for 5 minutes or when their EOD rate became
highly irregular (mean EOD rate / standard deviation < 2, values obtained for recent 10
minutes) these were taken as indicators for humane endpoint criteria. These criteria were not
fulfilled for any animal in our study.
3 / 17
In non-curarized fish, the EOD is driven by a descending spino-motor volley known as the
EOD motor-command signal (EODC). This descending command continues spontaneously
in curarized fish without, however, initiating an EOD. The EODC was recorded with a silver
wire bent around the fish's tail. The EODC then was used to trigger an artificial EOD stimulus
(Wavetek, Model 395) delivered at a delay after the first negative peak of the EODC (defined
as ªtime zeroº; t0). This delay was adjusted to match the fish's natural EODC/EOD interval,
which was recorded before curarisation (between 2.5 and 4.9 ms).
Phase-shifted (+10Ê or -10Ê) and unaltered (0Ê) artificial EODs were used as stimuli.
Phaseshifted EODs were constructed by retarding the phase angle of all positive frequencies of the
FFT phase spectrum of a pre-recorded EOD by a constant angle while the negative frequencies
were advanced by the same angle [
]. The time domain of these phase shifted EODs was
obtained through an inverse FFT. This ensured that all signals were similar in peak-to-peak
amplitude and power spectra. This is in contrast to natural capacitive objects, which induce
phase shifts in the range of 0Ê to -25Ê and additionally result in changes of the power spectrum
and the peak-to-peak amplitude. Thus, only B-cells of the mormyromasts should give
differential responses to the phase shifted signals used here . Stimuli were delivered through an
isolated symmetry amplifier (Elektronikwerkstatt, Bonn) to a pair of silver electrodes (exposed
diameter 2 mm, 1cm apart). This enabled us to stimulate small areas of the fish's skin. The
stimulation electrode was movable and positioned perpendicular to the skin. Measuring the
delivered EODs directly at the skin of the animal [
] showed that EODs remained similar in
their power spectra and peak-to-peak amplitudes, differing only in their peak ratios (Fig 1).
The motor-command signal was pre-amplified and passed through an amplifier/filter unit
(amplification x100, 10 Hz high-pass, 10 kHz low-pass, 50 Hz notch-filter, Elektronikwerkstatt,
Uni Bonn). Single cells in different layers of the ELL were recorded extracellularly using glass
microelectrodes (resistance 0.8±3.1 MO) filled with 3 M NaCl. Field potentials and single cell
recordings were amplified (DAM 80; WPI), band pass filtered (1Hz± 1kHz for field potentials;
300Hz-3kHz for single cell recordings) and notch filtered (HumBug; Quest Scientific). All
signals were displayed on an oscilloscope (DL 1540 CL; Yokogawa), digitised (10kHz; Power
1401; CED) and stored on a computer. The stereotyped electric-organ corollary discharge
(EOCD) evoked field potentials were used as landmarks for determining from which layer and
zone of the ELL we recorded [
After establishing a cell's receptive field, the stimulus electrode was positioned in the centre of
the receptive field at a lateral distance of 2 mm from the electroreceptor. To classify the cells,
we recorded their response to the EOCD alone as well as to a local stimulus that was
timelocked to the natural EOD timing (Fig 1C). This local stimulus was presented using at least five
different stimulus amplitudes. For subsequent tests the stimulus amplitude was set slightly
above a given cell's threshold. We distinguish between E- and I-cells: I-cells give a burst of
spikes in the absence of sensory stimuli and the number of spikes in the burst decreases with
increasing stimulus intensity, while the spike latency increased. E-cells give a burst of spikes to
a local sensory stimulus in the centre of their receptive field, but are mostly inactive in the
absence of sensory stimuli. An increase of stimulus amplitude causes an increase of spike rate
and a decrease in spike latency.
4 / 17
Fig 1. Confirmation of stimuli and schematic stimulus protocol. A-C: A. Power-spectra of the EODs used as
stimuli. For this analysis the EODs were measured next to the tip of the chin appendix of an animal. Single EODs for
the three conditions (0, + andÐ10Ê) are shown to the right. Note the differences in the positive-to-negative peak ratios
and the similarity of the power spectra. B. Exemplary local field potential recorded in the plexiform layer of the medial
zone of the ELL in the absence of sensory stimulation. The open triangle below the field potential recording indicates
the t0 reference, while the filled triangle indicates the time when the EOD would have occurred under natural
conditions, i.e. the time at which the artificial stimulus was presented. The schematic indicates the receptive field
centre of the encountered cell. C. Example of the stimulus protocol. From top to bottom: spikes (AP), EODs and phase
of the stimulus. For positive and negative phase shifts two consecutive phase shifts were presented. Each epoch lasted
for 30 seconds.
The effect of phase-shifted EODs was tested by stimulating with a basal EOD of zero phase
shift and EODs that were shifted by either +10Ê or -10Ê (referred to as F in the following).
Within a trial we switched between the basal and phase-shifted condition twice, starting and
ending with the basal condition. Each trial thus consisted of 5 epochs, each lasting 30 seconds,
with 2 phase-shifted epochs of either -10Ê or +10Ê (see Fig 1C). Negative phase shifts were
tested prior to positive phase shift. If recordings were stable, this protocol was repeated with
different EOD amplitudes.
To compare if responses within a trial differed between conditions, i.e. between basal and
phase-shifted epochs, we compared the spikes per EOD between epochs (Kruskal-Wallis test
followed by Dunn's post-hoc test, Matlab 11b). A nonparametric test was used as the number
of spikes per EOCD-cycle deviated from the normality assumption in some epochs
(Kolmogorov Smirnov test, SPSS 14). When a phase shifted epoch significantly differed from at least two
of the three zero-phase epochs within a trial we classified this as a reproducible
phase-sensitivity. In the Results section we report the number of all cells tested in a given condition followed
by the number of cells that were classified as reproducible (N all / N reproducible).
5 / 17
and stimulus condition (APF 1=n
To further analyse the effect of phase shifts per trial we obtained the mean number of spikes
per EOD for the un-shifted epochs (APbasal) and the mean number of spikes per EOD for the
phase-shifted epochs (APshifted). From these we calculated the mean number of spikes per EOD
APshifted, AP0 1=n
APbasal, with F being either +
or -10Ê). To quantify how responses differed between basal and phase-shifted conditions
within a trial, we calculated the mean rate-difference between the shifted and basal condition
DF APF AP0 . Mean rate-differences for positive shifts will be presented as Δ+10 and as
Δ−10 for negative phase-shifts. First spike latency was analysed in a similar manner and was
expressed relative to time zero (t0) of the EODC. To compare the effect of phase shifts between
I- and E-cells as well as between the DLZ and MZ zone, we used the difference of the mean
spike rates per group (one-way ANOVA followed by a Tukey's test, SPSS 14).
Different tracers were injected iontophoretically (4±5 μA DC current, 30 minutes, changing of
polarity every 5 minutes) at identified layers (ganglionic, plexiform or granular layer) of the
MZ or the DLZ during the electrophysiological experiments. Electrodes (resistance < 0.6 MO)
were filled with biocytin (4% in 3M NaCl), neurobiotin (4% in 3 M NaCl), or a fluorescent dye
(Fluoro-Ruby D-1817, 10 kDa, Invitrogen). After a survival time of 10±28 hours, the animals
were deeply anaesthetised (MS-222) and perfused with 2% paraformaldehyde and 2%
glutaraldehyde in phosphate buffer (0.1 M, pH 7.4).
Serial sections were cut on a vibratome (Vibratome1 1500, TSE systems; Leica 2000, Leica,
80 μm). Injections of biocytin were developed with ABC-complex (Vectastain1, ABC Kit,
PK4000; Vector Laboratories) and DAB (3,3'-Diaminobenzine) to reveal labelling. The sections
were mounted on glass slides (Thermo scientific Superfrost Plus™, Fischer Scientific, Illkirch,
France) and counterstained with neutral red. In some sections an additional fluorescent
Nisslstain was applied (NeuroTrace 530, Invitrogen). All fluorescent slices were mounted with
Vectashield1 (Vector Laboratories, Inc., H-1500).
As detailed in the introduction phase is represented in an ambiguous manner in the afferent
stream, yet Mormyrids can evaluate phase and amplitude independently [
]. However, no
evidence for convergence between both sensory streams has been found at the level of the ELL
in previous works [
]. As the medial and the dorsolateral maps of the ELL are
interconnected  and phase information is overrepresented in the part of the DLZ map that receives
input from the foveal chin appendix [
], this lack of interzonal processing in the ELL is
surprising. We here thus specifically investigated if interzonal processing of phase information
occurs in those regions of the ELL that receive input from the foveal areas. We report data
from 41 cells (medial zone = 29 cells, dorsolateral zone = 12 cells, see Table 1) and demonstrate
that cells in the medial map respond to phase-shifted stimuli.
Responses to phase-shifts in the DLZ
We first report results obtained in the DLZ, where neurones are known to be responsive to
phase-shifted stimuli. As expected, E-cells of the DLZ decreased the firing rate for positive
phase shifts (N = 6 / 9) and increased their firing rate in response to negative phase shifts
(N = 7 / 7, Table 1). This is shown for an exemplary cell in Fig 2, while the population data is
shown in Fig 6B.
6 / 17
Mean change in firing rate with respect to zero phase shifted firing rate for the DLZ and MZ maps separated between E- and I-cells and positive and negative phase
shifts. Data is presented for all cells followed by cells with significant and reproducible effects.
Fig 2. Example for an E-cell's response in the dorsolateral zone to +10Ê (A, B) and -10Ê (C, D) phase-shifts. This
cell responded with a reproducible de- (+10Ê) and increase (-10Ê) of its rate to the phase shifts, whereas first-spike
latency was not systematically altered. Here and in the following figures four panels (A-D) are shown. A, C. Raster
plots showing the change in spiking when switching from the undistorted (0Ê) to a phase-shifted (+ or -10Ê) EOD.
Responses to phase shifted EODs are visualized by the coloured background. Significant differences between
undistorted and phase-shifted conditions are indicated by the lines to the right (Kruskal-Wallis test with Dunn's
posthoc analysis, alpha = 0.05). The raster plots on the right side of panels A and C depict the duration of the EODC for the
corresponding raster plots. Note that EODC intervals were irregular and longer than the time at which spikes
occurred. For better visualisation the raster plots are thus shown to match the longest interval after time zero at which
spikes occurred in a given cell. B, D. Peri-stimulus time-histograms (PSTH) summarizing the data shown in A and C,
phase shifts are plotted in colour, undistorted EOD-data in black. See S1 File for data.
7 / 17
Fig 3. Example for an I-cell's response in the dorsolateral zone to +10Ê (left, A-B) and -10Ê (right, C-D)
phaseshifts. This cell responded reproducibly with an in- (+10Ê) or decreased (-10Ê) rate to the phase shifts, whereas
firstspike latency was not altered systematically. For the full legend to the panels, refer to Fig 2.
As expected, I-cells in the DLZ increased the firing rate in response to positive shifts
(N = 3 / 5) and decreased their rate in response to negative shifts (N = 3 / 5, see Figs 3 and 6
and Table 1).
In summary, our data on DLZ neurons corroborate published studies [
further indicates that our stimulation conditions were comparable to those used in these studies.
Responses to phase-shifts in the MZ
In the following we report results on phase-shifted stimuli in the medial zone, where previous
studies had not found evidence of phase-sensitivity. Three out of nine E-cells recorded in the
MZ increased their firing rate in response to positive phase shifts in a reproducible manner
(see Figs 4 and 6 and Table 1).
Negative phase shifts led to a reproducible reduction of the firing rate in three out of nine
E-cells tested (Figs 5 and 6). In I-cells, we found that eleven out of 20 cells decreased the firing
rate reproducibly when subjected to positive phase-shifts (Figs 5 and 6 and Table 1). When
stimulated with negative phase shifts, six out of these 20 cells increased their firing rates
Comparison between zones
A 2-way analysis of variance with the main effects of zone (MZ and DLZ) and cell-type (E- and
I-cells) was performed to compare the responses to phase shifts between zones and cell types.
This revealed the presence of disordinal main effects (F-ratios for +10Ê shifts: Zone: F(1,19) =
0.12, p = 0.73; cell-type: F(1,19) = 2.75, p = 0.11; F-ratios for -10Ê shifts: Zone: F(1,15) = 1.57,
8 / 17
Fig 4. Example for an E-cell's response in the medial zone to +10Ê (A, B) and -10Ê (C, D) phase-shifts. This cell
responded reproducibly with an in- (+10Ê) or decreased (-10Ê) rate to the phase shifts, whereas first-spike latency was
not altered significantly. For the full legend to the panels, refer to Fig 2. See S1 File for data.
p = .22; cell-type: F(1,15) = 4.59, p = 0.048) with significant interaction (F(1,19) = 30.68,
p < 0.001) for +10Ê phase shifts and F(1,15) = 45.38, p < 0.001 for -10Ê phase shifts). Hence
we focussed our analysis on the interaction effects, which were analysed using Tukey's HSD
post hoc test (Fig 6). As expected, E- and I-cells of the DLZ differed in their responses to phase
shifted EODs both for negative and positive phase-shifts with E-cells responding to negative
phase-shifts with an increase and I-cells with an decrease and vice versa for positive phase
shifts (Tukey's HSD, p < 0.001, Fig 6). The opposite effect was found for E- and I-cells in the
medial zone (Tukey's HSD, p < 0.05, Fig 6). A comparison between the zones confirmed that
a given cell-type of the MZ will respond in a manner opposite to the same cell-type's response
in the DLZ (see Fig 6). This indicates that the phase sensitivity of neurones in the medial zone
is not due to a direct (peripheral) input from the B-cells to this zone.
In order to investigate how phase-sensitivity is conveyed from the DLZ to the MZ, we traced
the connectivity between zones following tracer injections after either an electrophysiological
experiment (N = 9) or, in four cases, in fish specifically injected for this purpose. In all cases
injection sites were found in the zones targeted during the experiment, i.e., discrimination
between zones based on our macroscopic and physiological parameters was correct.
As expected [
], injections in either the DLZ or MZ labelled homotopical projections to
the other zone, i.e., the zones are interconnected and these connections maintain the
topography between the maps (Fig 7), as originally shown by Bell and colleagues [
]. At present
two cell types have been found to form interzonal connections, the Large multipolar
9 / 17
Fig 5. Example for an I-cell's response in the medial zone to +10Ê (left, A-B) and -10Ê (right, C-D) phase-shifts.
This cell responses reproducibly with a de- (10Ê) or an increased (-10Ê) rate to the phase shifts, whereas first-spike
latency was not altered (not shown). For the full legend to the panels, refer to Fig 2. See S1 File for data.
intermediate layer cells (LMI) projections [
] and the interzonal cell [
]. Both cell types
conserve topographic connections between zones where the interzonal cells terminate in the
superficial and deep granular layers, while the axons of the GABAergic LMI cells terminate in
the superficial granular layer. While we observed retrogradely labelled somata in the granular
layer (Fig 7A) as well as larger somata in the intermediate layers (arrowhead Fig 7C), stainings
were either too incomplete or dense to identify cell types. However, our data confirms the
presence of a substantial homotopic interzonal connection, which we propose is the most
parsimonious source of the phase-sensitivity described here for the cells of the MZ zone. Future
studies are required to determine the detailed source as well as the connectivity between the
zones to establish how the differential responses of E- and I-cells of the MZ and DLZ arise.
The independently evolved mormyroid and gymnotiforme weakly electric fish share a
topographically organized representation of different electrosensory features in the hindbrain. In
Gymnotiform fishes this parallel processing is achieved through differences in the
spatiotemporal tuning properties of secondary neurones, whereas in Mormyrids this parallel processing
already begins with two non-convergent sensory inputs from differently tuned
electroreceptors. The observation that Mormyrids can distinguish complex impedances has led to the
hypothesis that this may be based on a subtractive comparison of the phase and amplitude
pathway input. A pioneering study found no evidence for the required convergence of the two
sensory streams at the level of the ELL [
]. This is surprising given the significant interzonal
connections between the two maps in the ELL [
]. Processing of capacitive information
10 / 17
Fig 6. Summary graphs comparing the mean differences of I- and E-cells to phase-shifts in the medial and dorsolateral zone of the ELL.
Solid symbols show data from cells the DLZ and open symbols show data from cells in the MZ with circles indicating data from I-cells and
squares indicating data from E-cells. Responses to positive phase shifts are marked in red, while responses to negative shifts are marked in blue.
Error bars represent standard deviations. Note that similar cell types respond oppositely in both zones. See S1 File for data.
is particularly important for sensory input originating from the foveal chin appendix [
here investigated to which degree the parallel sensory streams are kept separate downstream
from the electroreceptors by specifically investigating the chin appendix region of the ELL.
Our results show that neurons from both the MZ and DLZ were affected by artificially
generated phase-shifted stimuli. Importantly, phase-shifted EODs had the same peak-to-peak
amplitude and power-spectral density as the non-shifted EODs (Fig 1) and only differed in the
P/N-ratios. While this would not occur under natural conditions, it enabled us to selectively
alter the response of B-cells. Any difference in neuronal response to shifted and non-shifted
EODs are hence attributable to a waveform-sensitivity of the recorded neuron.
Our results for neurons in the DLZ corroborate published data [
], showing the expected
decrease of the firing rate in I-cells and an increase in E-cells in response to negative phase
shifts. Our study unveiled a fundamental difference to prior studies with respect to the medial
zone. This zone receives only A-primary afferent input, and hence should not be responsive to
phase shifts below 30Ê [
]. While prior studies did not find an effect of phase-shifted stimuli
11 / 17
Fig 7. Anatomical data on interzonal connectivity. A. Low-magnification photomicrograph showing an ELL cross-section following
a biocytin injection in the medial part of the medial zone. The section has been processed following the DAB procedure and counter
stained with cresyl violet. The three zones are indicated on the slide and the different layers of the zones are indicated by the black lines
at the borders between the MZ and DLZ. Note that the commissural projection connects the medial and dorsolateral zone such that
the medial parts of the MZ connect with the lateral part of the DLZ. Likewise the lateral MZ is connected with the medial DLZ (see B,
C). As the dorso-ventral topography of the sensory surface is represented along the medio-lateral axis in the MZ and the lateral to
medial axis in the DLZ, this shows that somatotopically corresponding zones of the maps are connected. The higher magnification
inset on the left shows the area surrounded by the stippled line in the DLZ. The red arrows point towards two retrogradely labelled
somata in the granular layer. Abbreviations: deepf, deep fibre layer; gang, ganglionic layer; gran, granular layer; mol, molecular layer;
plex, plexiform layer. B. Photomicrograph showing an injection site in the lateral part of the MZ using neurobiotin 488 (shown in
green) and a fluorescent Nissl counterstain (blue). C. Interzonal projection to the DLZ originating from the injection shown in B. Note
that in addition to the strong labelling of fibres some weakly stained large somata are present in the interzonal layer. The schematic
inset in A and B shows a cross-section of the medulla with the injection site and the region where the close-up were taken indicated in
green. deep, deep fibre layer; ggl, ganglionic layer, gran, granular layer, inter, intermediate layer; plex, plexiform layer.
12 / 17
for neurons of the MZ that process sensory input form the trunk [
], our recordings of
MZ-neurons that receive sensory input from the chin appendix showed that about half of all
cells responded to phase-shifted stimuli well below 30Ê. We specifically focused on the chin
appendix because this appears to be a sensory fovea devoted to impedance analysis . We
therefore expected that evidence for interzonal processing should be obtained in this part of
the ELL most clearly. A second modification with respect to the earlier studies concerns the
timing of the stimuli. We matched the timing of the stimuli to match the EOD timing
measured prior to each experiment. The delay between the motor-command signal and the EOD
varied between 2.4 and 4.9 ms in different fish, whereas the previous studies used a fixed delay
of 5 ms. As sensory processing in the ELL largely depends on the timing of the interaction of a
centrally originating corollary input to the ELL with the sensory input [
], this discrepancy
likely has a considerable influence on the observed responses. It remains to be tested whether
similar results might be obtained for neurones of the MZ that process sensory input from the
We suggest that the phase-sensitivity of the MZ is based on interzonal input. The two
known cell types known to make interzonal connections both will be sensitive to a mismatched
timing of corollary and sensory input [
]. Thus, it remains to be shown if the previous lack of
evidence for phase-sensitivity is specific to the trunk regions of ELL, or rather related to the
timing mismatch in earlier studies. For the responses of the DLZ the cause of the differential
responses of E- and I-cells is the connectivity within this zone: negative phase shifts lead to an
increased activity at reduced latency in the mormyromast B-cell primary afferents. These
terminate on granular cells that make inhibitory synapses with I-cells (e.g. LG and MG1 cells)
and are assumed to make excitatory synapses with E-cells (e.g. LF- and MG2 cells) [
Phaseshifts thus increase the inhibition of I-cells and increase the excitation of E-cells. This is in
agreement with both our and the aforementioned studies. Future studies are required to
address the cells and connectivity between the two zones. Our labelling confirmed the presence
of reciprocal interzonal connections. Hence it remains to be investigated if and how the medial
zone influences responses of the dorsolateral zone.
We here have reported results using artificial stimuli that were designed to deliver very
local sensory activation at the periphery. The efferent neurones of the ELL are known to have
centre-surround organisation [
], and it remains to be shown if centre and surround
contribute differently to the phase sensitivity of neurones in the medial zone. Furthermore, under
natural conditions, the sensory input consists of a global stimulation of the full array of
electroreceptors. In a subset of cells we also tested the effect of global stimuli. However, as we only
were able to do so in a total of eleven cells, of which 3 responded reproducibly to local shifts of
the stimulus phase, we could not systematically analyse this data here. It should be noted
however, that two neurones of the MZ that were classified as phase-sensitive also responded to
globally shifted stimuli. His may suggest that the mediated phase-sensitivity is dominated by a
cells centre, assuming that our local stimuli did selectively stimulate the centre. It has been
shown that responses to global stimulation can differ considerably from responses to local
stimuli such as the ones used in our present study . If future studies confirm that phase
sensitivity in the medial zone does not differ between local and global stimulation, this may further
suggest that feedback from higher order electrosensory centres like the preeminential nucleus
], are not involved in mediating this phase sensitivity.
In summary, our data are a first step to understand how weakly electric Mormyrid fish
analyse their comparatively simple sensory environment through parallel processing. While we
did not find ªcapacitanceº encoding units in the ELL, we have shown that the parallel
information is used to enhance the contrast between the two sensory streams at higher stages. Merging
of these optimized ELL outputs most likely takes place at the torus semicircularis. To better
13 / 17
understand the function of the two maps, future studies should aim to reveal the
spatiotemporal properties of the dorsolateral zone to investigate if feature-optimized encoding schemes as
reported for the three maps in Gymnotiform weakly electric fish (e.g., 50±52) exist in
Mormyrid parallel maps as well.
S1 File. Data tables to reproduce Figs 2±6. The supporting information is provided in form
of nine separate files compressed in a single RAR-archive. Files named Fig2a.txt to Fig5b.txt
contain the data to reproduce figures 2±5, while the file Fig6.txt contains the data to reproduce
the data shown in figure six for all cells as well as the cells that responded in a reproducible
manner to phase shifts. In all files the first row indicates what kind of data is being shown in
the columns. Data in columns named AP1 to AP14 (Fig2a.txt to Fig5b.txt) show the latency of
spikes in response to the EOD mimic in seconds.
All authors had full access to the data in the study and take responsibility for the integrity of
the data and the accuracy of the data analysis. Study concept and design: J.E. Acquisition of
data: S.F. Analysis and interpretation of data: S.F., J.E., K.G and G.v.d.E. Drafting of the
manuscript: S.F and J.E. Statistical analysis: S.F. and J.E. Administrative and material support: K.G.
and G.v.d.E. Study supervision: J.E, K.G. and G.v.d.E. Technical support for histology and
microscopy in Gif-sur-Yvette: Guillaume Hucher. The authors would like to thank two
anonymous reviewers for their constructive support.
Conceptualization: Sylvia Fechner, Gerhard von der Emde, Jacob Engelmann.
Data curation: Jacob Engelmann.
Formal analysis: Sylvia Fechner, Jacob Engelmann.
Funding acquisition: Kirsty Grant, Gerhard von der Emde, Jacob Engelmann.
Investigation: Sylvia Fechner, Kirsty Grant, Jacob Engelmann.
Methodology: Kirsty Grant, Gerhard von der Emde, Jacob Engelmann.
Project administration: Gerhard von der Emde, Jacob Engelmann.
Resources: Gerhard von der Emde, Jacob Engelmann.
Software: Jacob Engelmann.
Supervision: Gerhard von der Emde, Jacob Engelmann.
Validation: Sylvia Fechner, Kirsty Grant, Gerhard von der Emde, Jacob Engelmann.
Visualization: Sylvia Fechner, Kirsty Grant, Jacob Engelmann.
Writing ± original draft: Sylvia Fechner, Kirsty Grant, Gerhard von der Emde, Jacob
Writing ± review & editing: Sylvia Fechner, Gerhard von der Emde, Jacob Engelmann.
14 / 17
15 / 17
16 / 17
1. Kaas J . Topographic maps are fundamental to sensory processing . Brain Res Bull [Internet] . 1997 ; 44 ( 2 ): 107 ± 12 . PMID: 9292198
1997 Jan; 44 ( 2 ): 113 ± 6 . PMID: 9292199
3. van Hemmen JL. The map in your head: How does the brain represent the outside world ? Vol. 3 , ChemPhysChem. 2002 . p. 291 ± 8 .
4. Brown WM , BaÈcker A. Optimal neuronal tuning for finite stimulus spaces . Neural Comput [Internet] . 2006 Jul; 18 ( 7 ): 1511 ± 26 . https://doi.org/10.1162/neco. 2006 . 18 .7.1511 PMID: 16764512
5. Schreiner CE , Winer JA . Auditory cortex mapmaking: principles, projections, and plasticity . Neuron [Internet] . 2007 ; 56 ( 2 ): 356 ± 65 . https://doi.org/10.1016/j.neuron. 2007 . 10 .013 PMID: 17964251
Winer JA , Miller LM , Lee CC , Schreiner CE . Auditory thalamocortical transformation: structure and function . Trends Neurosci [Internet] . 2005 May; 28 ( 5 ): 255 ± 63 . https://doi.org/10.1016/j.tins. 2005 . 03 .
009 PMID: 15866200
7. Lee CC , Schreiner CE , Imaizumi K , Winer JA . Tonotopic and heterotopic projection systems in physiologically defined auditory cortex . Neuroscience [Internet] . 2004 Jan; 128 ( 4 ): 871 ± 87 . https://doi.org/10. 1016/j.neuroscience. 2004 . 06 .062 PMID: 15464293
8. Graziano MS , A TN . Rethinking cortical organization: moving away from discrete areas arranged in hierarchies . Neuroscientist [Internet] . 2007 Apr; 13 ( 2 ): 138 ± 47 . https://doi.org/10.1177/1073858406295918 PMID: 17404374
9. Lissmann H , Machin K. The mechanism of object location in Gymnarchus niloticus and similar fish . J Exp Biol [Internet] . 1958 ; 35 : 451 ± 86 .
10. Krahe R , Maler L. Neural maps in the electrosensory system of weakly electric fish . Curr Opin Neurobiol [Internet] . 2014 Feb; 24 : 13 ± 21 . https://doi.org/10.1016/j.conb. 2013 . 08 .013 PMID: 24492073
11. Braford MR , McCormick C . Brain organization in teleost fishes: lessons from the electrosense . J Comp Physiol A . 1993 ; 173 : 704 ± 7 .
12. Metzner W , Juranek J. A sensory brain map for each behavior ? Proc Natl Acad Sci USA [Internet] . 1997 ; 94 (December): 14798 ± 803 .
13. Shumway C . Multiple electrosensory maps in the medulla of weakly electric gymnotiform fish. I. Physiological differences . J Neurosci [Internet] . 1989 ; 9 (December): 4388 ± 99 .
14. Kaas J . Why does the brain have so many visual areas? J Cogn Neurosci . 1989 ; 1 ( 2 ): 121 ± 35 . https:// doi.org/10.1162/jocn. 1989 . 1 .2.121 PMID: 23968461
15. Barlow H . Why have multiple cortical areas? Vision Res [Internet]. 1986 ; 26 ( 1 ): 81 ± 90 . PMID: 3716216
16. Kaas J. The segregation of function in the nervous system: Why do sensory systems have so many subdivisions . In: Neef WP, editor. Contributions to sensory physiology. New York: Academic Press; 1982 . p. 200 ± 40 .
17. Vonderschen K , Chacron MJ . Sparse and dense coding of natural stimuli by distinct midbrain neuron subpopulations in weakly electric fish . 2011 ; 3102 ± 18 .
18. McGillivray P , Vonderschen K , Fortune ES , Chacron MJ . Parallel coding of first- and second-order stimulus attributes by midbrain electrosensory neurons . J Neurosci [Internet] . 2012 Apr 18 ; 32 ( 16 ): 5510 ± 24 . https://doi.org/10.1523/JNEUROSCI.0478- 12 . 2012 PMID: 22514313
19. Khosravi-Hashemi N , Chacron M. Bursts and Isolated Spikes Code for Opposite Movement Directions in Midbrain Electrosensory Neurons . PLoS One [Internet] . 2012 ; 7 ( 6 ):e40339. https://doi.org/10.1371/ journal.pone. 0040339 PMID: 22768279
20. von der Emde G , Bleckmann H . Waveform tuning of electroreceptor cells in the weakly electric fish, Gnathonemus petersii . J Comp Physiol A [Internet] . 1997 ; 511 ± 24 .
21. Szabo T , WersaÈll J. Ultrastructure of an electroreceptor (mormyromast) in a mormyrid fish, Gnathonemus petersii . II. J Ultrastruct Res [Internet] . 1970 ; 490 ( 5 ): 473 ± 90 .
22. Szabo T , Hagiwara S. A latency-change mechanism involved in sensory coding of electric fish . Physiol Behav [Internet] . 1967 ; 2 ( 4 ): 331 ± 335 .
23. Bell CC . Mormyromast electroreceptor organs and their afferent fibers in mormyrid fish . III. Physiological differences between two morphological types of fibers . J Neurophysiol [Internet] . 1990 ; 63 ( 2 ): 319 ± 32 . https://doi.org/10.1152/jn. 1990 . 63 .2.319 PMID: 2313348
24. von der Emde G , Bleckmann H. Differential responses of two types of electroreceptive afferents to signal distortions may permit capacitance measurement in a weakly electric fish, Gnathonemus . J Comp Physiol A [Internet]. 1992 ; 171 ( 5 ): 683 ± 94 .
25. von der Emde G , Bleckmann H . Extreme phase sensitivity of afferents which innervate mormyromast electroreceptors . Naturwissenschaften [Internet] . 1992 ; 133 : 131 ± 3 .
26. von der Emde G. Discrimination of objects through electrolocation in the weakly electric fish, Gnathonemus petersii . J Comp Physiol A [Internet] . 1990 Aug; 167 ( 3 ): 413 ± 21 .
27. von der Emde G , Ronacher B . Perception of electric properties of objects in electrolocating weakly electric fish: two-dimensional similarity scaling reveals a City-Block metric . J Comp Physiol A [Internet] . 1994 ; 801 ± 12 .
28. von der Emde G. The sensing of electrical capacitances by weakly electric Mormyrid fish: effects of water conductivity . J Exp Biol [Internet] . 1993 Aug 1 ; 173 ( 1 ): 157 ± 73 .
29. Bell CC , Maler L . Central Neuroanatomy of Electrosensory Systems in Fish . In: Bullock TH , Hopkins CD , Popper AN , Fay RR , editors. Electroreception [Internet] . Springer H. New York: Springer New York; 2005 . p. 68 ± 111 .
30. Bell CC , Finger TE , Russell CJ . Central connections of the posterior lateral line lobe in mormyrid fish . Exp Brain Res [Internet] . 1981 ; 42 ( 1 ):9± 22 . PMID: 6163655
31. von der Emde G , Bell CC . Responses of cells in the mormyrid electrosensory lobe to EODs with distorted waveforms: implications for capacitance detection . J Comp Physiol A [Internet] . 1994 ; 83 ± 93 .
32. Bacelo J , Engelmann J , Hollmann M , von der Emde G , Grant K. Functional foveae in an electrosensory system . J Comp Neurol [Internet] . 2008 Nov 20 ; 511 ( 3 ): 342 ± 59 . https://doi.org/10.1002/cne.21843 PMID: 18803238
33. Bell CC , Grant K , Serrier J , Bell CC , Grant K , Serrier J . Sensory processing and corollary discharge effects in the mormyromast regions of the mormyrid electrosensory lobe. I. Field potentials, cellular activity in associated structures . J Neurophysiol [Internet] . 1992 Sep; 68 ( 68 ): 843 ± 58 .
34. Goenechea L , von der Emde G. Responses of neurons in the electrosensory lateral line lobe of the weakly electric fish Gnathonemus petersii to simple and complex electrosensory stimuli . J Comp Physiol A [Internet] . 2004 Nov; 190 ( 11 ): 907 ± 22 .
35. Hitschfeld EM , Stamper SA , Vonderschen K , Fortune ES , Chacron MJ . Effects of restraint and immobilization on electrosensory behaviors of weakly electric fish . ILAR J . 2009 ; 50 ( 4 ): 361 ± 72 . PMID: 19949252
36. Hopkins C , Bass A. Temporal coding of species recognition signals in an electric fish . Science [Internet] . 1981 ; 212 : 85 ± 8 . PMID: 7209524
37. Heiligenberg W , Altes R . Phase sensitivity in electroreception . Science [Internet] . 1978 ; 199 ( 4332 ): 1001 ± 4 . PMID: 622577
38. Engelmann J , Bacelo J , van den Burg E , Grant K. Sensory and motor effects of etomidate anesthesia . J Neurophysiol [Internet] . 2006 Feb; 95 ( 2 ): 1231 ± 43 . https://doi.org/10.1152/jn.00405. 2005 PMID: 16267119
39. Meek J , Hafmans TG , Han V , Bell CC , Grant K. Myelinated dendrites in the mormyrid electrosensory lobe . J Comp Neurol [Internet] . 2001 Mar 12 ; 431 ( 3 ): 255 ± 75 . PMID: 11170004
40. Finger T , Bell C , Russell C. Electrosensory pathways to the valvula cerebelli in mormyrid fish . Exp Brain Res [Internet] . 1981 ;( 42): 23 ± 33 . PMID: 6163654
41. Bell CC , Finger T , Russell C . Central connections of the posterior lateral line lobe in mormyrid fish . Exp Brain Res [Internet] . 1981 ; ( 42):9±22 . PMID: 6163655
42. Bell CC , Russell C . Termination of electroreceptor and mechanical lateral line afferents in the mormyrid acousticolateral area . J Comp Neurol [Internet] . 1978 Dec 1 ; 182 ( 3 ): 367 ± 82 . https://doi.org/10.1002/ cne.901820302 PMID: 721966
43. Meek J , Hafmans TGM , Han V , Bell CC , Grant K. Myelinated dendrites in the mormyrid electrosensory lobe . J Comp Neurol . 2001 ; 431 ( 3 ): 255 ± 75 . PMID: 11170004
44. Mohr C , Roberts PD , Bell CC . The mormyromast region of the mormyrid electrosensory lobe. I. Responses to corollary discharge and electrosensory stimuli . J Neurophysiol . 2003 ; 90 ( 2 ): 1193 ± 210 . https://doi.org/10.1152/jn.00211. 2003 PMID: 12904505
45. Meek J , Kirchberg G , Grant K , von der Emde G. Dye coupling without gap junctions suggests excitatory connections of gamma-aminobutyric acidergic neurons . J Comp Neurol [Internet] . 2004 Jan 6 ; 468 ( 2 ): 151 ± 64 . https://doi.org/10.1002/cne.10951 PMID: 14648676
46. Bell CC , Szabo T. Electroreception in mormyrid fish. Central anatomy . In: Heiligenberg W, Bullock TH , editors. Electroreception . New York: John Wiley & Sons; 1986 . p. 375 ± 421 .
47. Sawtell NB , Williams A , Bell CC . From sparks to spikes: Information processing in the electrosensory systems of fish . Curr Opin Neurobiol . 2005 ; 15 ( 4 ): 437 ± 43 . https://doi.org/10.1016/j.conb. 2005 . 06 .006 PMID: 16009545
48. Meek J , Grant K , Bell C . Structural organization of the mormyrid electrosensory lateral line lobe . J Exp Biol [Internet] . 1999 May; 202(Pt 10 ): 1291 ± 300 . PMID: 10210669
49. Bell C , Caputi A , Grant K. Physiology and plasticity of morphologically identified cells in the mormyrid electrosensory lobe . J Neurosci [Internet] . 1997 ; 17 ( 16 ): 6409 ± 23 . PMID: 9236249
50. Metzen MG , Engelmann J , Bacelo J , Grant K , von der Emde G. Receptive field properties of neurons in the electrosensory lateral line lobe of the weakly electric fish, Gnathonemus petersii . J Comp Physiol A Neuroethol Sens Neural Behav Physiol [Internet] . 2008 Dec; 194 ( 12 ): 1063 ± 75 . https://doi.org/10.1007/ s00359-008 -0377-4 PMID: 18855000
51. Goenechea L , Von Der Emde G . Responses of neurons in the electrosensory lateral line lobe of the weakly electric fish Gnathonemus petersii to simple and complex electrosensory stimuli . J Comp Physiol A Neuroethol Sensory , Neural, Behav Physiol . 2004 ; 190 ( 11 ): 907 ± 22 .
52. von der Emde G , Bell CC . Nucleus preeminentialis of mormyrid fish, a center for recurrent electrosensory feedback. I. Electrosensory and corollary discharge responses . J Neurophysiol . 1996 ; 76 : 1581 ± 96 . https://doi.org/10.1152/jn. 1996 . 76 .3.1581 PMID: 8890278