Electric interactions through chirping behavior in the weakly electric fish, Apteronotus leptorhynchus

The weakly electric fish Apteronotus leptorhynchus produces wave-like electric organ discharges distinguished by a high degree of regularity. Transient amplitude and frequency modulations (“chirps”) can be evoked in males by stimulation with the electric field of a conspecific. During these interactions, the males examined in this study produced six types of chirps, including two novel ones. Stimulation of a test fish with a conspecific at various distances showed that two electrically interacting fish must be within 10 cm of each other to evoke chirping behavior in the neighboring fish. The chirp rate of all but one chirp type elicited by the neighboring fish was found to be negatively correlated with the absolute value of the frequency difference between the two interacting fish, but independent of the sign of this difference. Correlation analysis of the instantaneous rates of chirp occurrence revealed two modes of interactions characterized by reciprocal stimulation and reciprocal inhibition. Further analysis of the temporal relationship between the chirps generated by the two fish during electric interactions showed that the chirps generated by one individual follow the chirps of the other with a short latency of approximately 500–1000 ms. We hypothesize that this “echo response” serves a communicatory function.     

In vitro studies of closed-loop feedback and electrosensory processing in Apteronotus leptorhynchus

Electrosensory systems comprise extensive feedback pathways. It is also well known that these pathways exhibit synaptic plasticity on a wide-range of time scales. Recent in vitro brain slice studies have characterized synaptic plasticity in the two main feedback pathways to the electrosensory lateral line lobe (ELL), a primary electrosensory nucleus in Apteronotus leptorhynchus. Currently-used slice preparations, involving networks in open-loop conditions, allow feedback inputs to be studied in isolation, a critical step in determining their synaptic properties. However, to fully understand electrosensory processing, we must understand how dynamic feedback modulates neuronal responses under closed-loop conditions. To bridge the gap between current in vitro approaches and more complex in vivo work, we present two new in vitro approaches for studying the roles of closed-loop feedback in electrosensory processing. The first involves a hybrid-network approach using dynamic clamp, and the second involves a new slice preparation that preserves one of the feedback pathways to ELL in a closed-loop condition.     

Ontogeny of the electric organ discharge and the electric organ in the weakly electric pulse fish Brachyhypopomus pinnicaudatus (Hypopomidae, Gymnotiformes)

I recorded the electric organ discharges (EODs) of 331 immature Brachyhypopomus pinnicaudatus 6–88 mm long. Larvae produced head-positive pulses 1.3 ms long at 7 mm (6 days) and added a second, small head-negative phase at 12 mm. Both phases shortened duration and increased amplitude during growth. Relative to the whole EOD, the negative phase increased duration until 22 mm and amplitude until 37 mm. Fish above 37 mm produced a “symmetric” EOD like that of adult females. I stained cleared fish with Sudan black, or fluorescently labeled serial sections with anti-desmin (electric organ) or anti-myosin (muscle). From day 6 onward, a single electric organ was found at the ventral margin of the hypaxial muscle. Electrocytes were initially cylindrical, overlapping, and stalk-less, but later shortened along the rostrocaudal axis, separated into rows, and formed caudal stalks. This differentiation started in the posterior electric organ in 12-mm fish and was complete in the anterior region of fish with “symmetric” EODs. The lack of a distinct “larval” electric organ in this pulse-type species weakens the hypothesis that all gymnotiforms develop both a temporary (larval) and a permanent (adult) electric organ. Accepted: 1 March 1997     

Social interaction and cortisol treatment increase cell addition and radial glia fiber density in the diencephalic periventricular zone of adult electric fish, Apteronotus leptorhynchus

In electric fish, Apteronotus leptorhynchus, both long-term social interaction and cortisol treatment potentiates chirping, an electrocommunication behavior that functions in aggression. Chirping is controlled by the diencephalic prepacemaker nucleus (PPn-C) located just lateral to the ventricle. Cells born in adult proliferative zones such as the periventricular zone (PVZ) can migrate along radial glial fibers to other brain regions, including the PPn-C. We examined whether social interactions or cortisol treatment influenced cell addition and radial glia fiber formation by (1) pairing fish (4 or 7 days) or (2) implanting fish with cortisol (7 or 14 days). Adult fish were injected with bromodeoxyuridine 3 days before sacrifice to mark cells that were recently added. Other fish were sacrificed after 1 or 7 days of treatment to examine vimentin immunoreactivity (IR), a measure of radial glial fiber density. Paired fish had more cell addition than isolated fish at 7 days, coinciding temporally with the onset of socially induced increase in chirping behavior. Paired fish also had higher vimentin IR at 1 and 7 days. For both cell addition and vimentin IR, the effect was regionally specific, increasing in the PVZ adjacent to the PPn-C, but not in surrounding regions. Cortisol increased cell addition at 7 days, correlating with the onset of cortisol-induced changes in chirping, and in a regionally specific manner. Cortisol for 14 days increased cell addition, and cortisol for 7 days increased vimentin IR but in a regionally non-specific manner. The correlation between treatment-induced changes in chirping and regionally specific increases in cell addition, and radial glial fiber formation suggests a causal relationship between such behavioral and brain plasticity in adults, but this hypothesis will require further testing.     

Glucocorticoid receptor blockade inhibits brain cell addition and aggressive signaling in electric fish, Apteronotus leptorhynchus

When animals are under stress, glucocorticoids commonly inhibit adult neurogenesis by acting through glucocorticoid receptors (GRs). However, in some cases, conditions that elevate glucocorticoids promote adult neurogenesis, and the role of glucocorticoid receptors in these circumstances is not well understood. We examined the involvement of GRs in social enhancement of brain cell addition and aggressive signaling in electric fish, Apteronotus leptorhynchus. In this species, long-term social interaction simultaneously elevates plasma cortisol, enhances brain cell addition and increases production of aggressive electrocommunication signals (“chirps”). We implanted isolated and paired fish with capsules containing nothing (controls) or the GR antagonist, RU486, recorded chirp production and locomotion for 7 d, and measured the density of newborn cells in the periventricular zone. Compared to isolated controls, paired controls showed elevated chirping in two phases: much higher chirp rates in the first 5 h and moderately higher nocturnal rates thereafter. Treating paired fish with RU486 reduced chirp rates in both phases to those of isolated fish, demonstrating that GR activation is crucial for socially induced chirping. Neither RU486 nor social interaction affected locomotion. RU486 treatment to paired fish had a partial effect on cell addition: paired RU486 fish had less cell addition than paired control fish but more than isolated fish. This suggests that cortisol activation of GRs contributes to social enhancement of cell addition but works in parallel with another GR-independent mechanism. RU486 also reduced cell addition in isolated fish, indicating that GRs participate in the regulation of cell addition even when cortisol levels are low.     

Gradual frequency rises in interacting black ghost knifefish,Apteronotus albifrons

The present paper highlights the relationship between social status and production of gradual frequency rises in interacting Apteronotus albifrons. The gradual frequency rise production was mathematically inferred and a discrete classification deliberately avoided. The results showed little gradual frequency rise production before the hierarchy settlement. Afterwards, only the dominant fish kept this gradual frequency rise production at low levels, while the subdominant fish drastically increased it in all following interaction contexts. The hypothesis of gradual frequency rises being involved in communication as submissive signals was thus strengthened.Abbreviations EOD electric organ discharge - FM frequency modulation - GFR gradual frequency rise - IQR interquartile range - JAR jamming avoidance response     

Plasticity of the electric organ discharge waveform of the electric fish Brachyhypopomus pinnicaudatus I. Quantification of day-night changes

The electric organ discharge of the gymnotiform fish Brachyhypopomus pinnicaudatus is a biphasic waveform. The female's electric organ discharge is nearly symmetric but males produce a longer second phase than first phase. In this study, infrared-sensitive video cameras monitored the position of unrestrained fish, facilitating precise measurement of electric organ discharge duration and amplitude every 2 h for 24 h. Males (n=27) increased electric organ discharge duration by 37 ± 12% and amplitude by 24 ± 9% at night and decreased it during the day. In contrast, females (n=8) exhibited only minor electric organ discharge variation over time. Most of a male's increase occurred rapidly within the first 2–3 h of darkness. Electric organ discharge values gradually diminished during the second half of the dark period and into the next morning. Modulation of the second phase of the biphasic electric organ discharge produced most of the duration change in males, but both phases changed amplitude by similar amounts. Turning the lights off at mid-day triggered an immediate increase in electric organ discharge, suggesting modification of existing ion channels in the electric organ, rather than altered genomic expression. Exaggeration of electric organ discharge sex differences implies a social function. Daily reduction of duration and amplitude may reduce predation risk or energy expenditure. Accepted: 12 September 1998     

Profile of micronucleus frequencies and nuclear abnormalities in different species of electric fishes (Gymnotiformes) from the Eastern Amazon

The frequency of spontaneous micronucleus (MN) formation in fish species needs to be determined to evaluate their usefulness for genotoxic biomonitoring. The definition of a good bioindicator takes into account the current knowledge of its metabolic traits as well as other factors including its feeding behavior and relationship to the environment. In this study, we compared the basal frequencies of micronucleated erythrocytes and nuclear abnormalities (NA) among different species of the fish Order Gymnotiformes (Rhamphichthys marmoratus, Steatogenys elegans, Sternopygus macrurus, Parapteronotus hasemani, Gymnotus mamiraua, Gymnotus arapaima, Brachyhypopomus beebei, Brachyhypopomus n. sp. BENN) sampled in several localities of the Eastern Amazon. A baseline of MN and NA frequency in these fish was determined, enabling the identification of potentially useful species as models for genotoxicity studies. Only one impacted sample collected at a site in the River Caripetuba showed a significant number of NAs, which may be due to the release of wastewater by neighbouring mining industries and by the burnt fuel released by the small boats used by a local community. Our results may provide support for further studies in areas of the Eastern Amazon affected by mining, deforestation and other anthropogenic activities.     

Phase and amplitude maps of the electric organ discharge of the weakly electric fish,Apteronotus leptorhynchus

Summary The electric organ discharge (EOD) potential was mapped on the skin and midplane of several Apteronotus leptorhynchus. The frequency components of the EOD on the surface of the fish have extremely stable amplitude and phase. However, the waveform varies considerably with different positions on the body surface. Peaks and zero crossings of the potential propagate along the fish's body, and there is no point where the potential is always zero. The EOD differs significantly from a sinusoid over at least one third of the body and tail. A qualitative comparison between fish showed that each individual had a unique spatiotemporal pattern of the EOD potential on its body.The potential waveforms have been assembled into high temporal and spatial resolution maps which show the dynamics of the EOD. Animation sequences and Macintosh software are available by anonymous ftp (mordor.cns.caltech.edu; cd/pub/ElectricFish).We interpret the EOD maps in terms of ramifications on electric organ control and electroreception. The electrocytes comprising the electric organ do not all fire in unison, indicating that the command pathway is not synchronized overall. The maps suggest that electroreceptors in different regions fulfill different computational roles in electroreception. Receptor mechanisms may exist to make use of the phase information or harmonic content of the EOD, so that both spatial and temporal patterns could contribute information useful for electrolocation and communication.Abbreviations EOD electric organ discharge - EO electric organ - CV coefficient of variance     

Social competition affects electric signal plasticity and steroid levels in the gymnotiform fish Brachyhypopomus gauderio

Sexually-selected communication signals can be used by competing males to settle contests without incurring the costs of fighting. Steroid regulation of these signals can render them as reliable indicators of a male's physiological state. We investigated how plasticity in electrocommunication signals is driven by social competition for mates, mediated by steroid hormones, and subject to the effects of past social experience. We measured the electric waveform's amplitude and duration and steroid hormone levels of male gymnotiform electric fish (Brachyhypopomus gauderio) following week-long periods of social isolation, and low or high social competition. To quantify the effect of social history on the modulation of the electric signal, six groups of six males experienced all three social conditions but in different order. We found that males differentially modulate their electric signals depending on the order they experienced these conditions. Thus, past social interactions affect both present and future social electric signals. Cortisol levels and the amplitude of the electric signal appeared to track the intensity of competition, while androgen levels and the duration of the electric signal only responded to the presence (low and high competition) or absence (isolation) of a social environment (low and high androgens respectively). In addition, cortisol levels were related to the body size of the males at high social competition. Taken together, these findings suggest that the capacity of males to modulate their signals in response to social competition is regulated by steroids.     

Electric organ discharge frequency and plasma sex steroid levels during gonadal recrudescence in a natural population of the weakly electric fish Sternopygus macrurus

1. Sternopygus macrurus were collected in Venezuela during the period of gonadal recrudescence in early or late dry season. Electric organ discharge (EOD) frequencies were recorded, blood samples were taken for analysis of steroid titers, and gonads were taken for determination of reproductive condition. 2. Mean EOD frequencies were significantly lower in males than in females in all samples. EOD frequency was inversely correlated with body length in males in late, but not early, dry season, and these parameters were never correlated in females. 3. Plasma levels of testosterone (T) and 11-ketotestosterone (11-KT), but not estradiol-17 beta (E2), were inversely correlated with EOD frequency in males. No 11-KT was observed in plasma of females, and plasma levels of T and E2 in females were comparable to those of males. Neither T nor E2 were correlated with EOD frequency in females. 4. Testes collected in late dry season were more mature than those from early dry season; androgen levels and EOD frequency were correlated with testicular maturity. Ovaries collected in early dry season were immature, while those from late dry season were more mature. There was no relationship between EOD frequency and stage of ovarian development. 5. These results suggest that plasma androgens modulate EOD frequency in males during the reproductive season and that plasma E2 has little relationship to EOD frequency in either sex.     

Time coding in the midbrain of mormyrid electric fish. I. Physiology and anatomy of cells in the nucleus exterolateralis pars anterior

In mormyrid electric fish, species-specific electric organ discharge waveforms are thought to be analyzed by the Knollenorgan electroreceptor subsystem. The midbrain anterior and posterior exterolateral nuclei (ELa and ELp) are thought to be the sites of this analysis. This paper is an electrophysiological study of the properties of the neurons in ELa. We recorded intracellularly from three classes of cells within ELa: the afferent axons from the nucleus of the electrosensory lateral line lobe (NELL), the large interstitial cells of ELa and an unidentified cell type. The large cells and the NELL axons were identified by intracellular injection of biocytin and are physiologically similar. Cells in ELa responded to square pulse stimuli with one or more time-locked action potentials with 2.8–3.0 ms latency. Both large cells and NELL axons arborized extensively in ELa and contacted numerous small cells. Based on the pattern of arborizations, we constructed a counter- current flow model of temporal coding by the small cells of ELa. We postulate that individual small cells are not selectively tuned for specific stimulus durations, but rather, the firing patterns of groups of small cells must be analyzed by neurons further up in the sensory hierarchy to determine the stimulus duration. Accepted: 25 June 1997     

Spatial distribution of the medullary command signal within the electric organ of Gymnotus carapo

The pacemaker nucleus of Gymnotus carapo contains two types of neurons: pacemaker cells which set up the frequency of the electric organ discharge (EOD) and relay cells which convey the command signal to the spinal cord. Direct activation of a single relay cell provides enough excitation to discharge a pool of spinal electromotor neurons and electrocytes, generating a small EOD (unit EOD). Different relay cells generate unit EODs of variable size and waveform, indicating the involvement of different groups of electrocytes. A special technique of EOD recording (multiple air-gap) was combined with intracellular stimulation of relay cells to study the spatial distribution within the electric organ (EO) of the command signal arising from different relay cells. Three types of relay cells could be identified: type I commanding the rostral 10% of the EO, type II which distribute their command all along the EO and type III driving the caudal 30%. Waveform analysis of unit EODs indicates that doubly innervated electrocytes which are the most relevant for attaining the specific EOD waveform, receive a favored command from the pacemaker nucleus.Abbreviations CV conduction velocity - EMF electromotive force - EMN electromotor neuron - EO electric organ - EOD electric organ discharge - PN pacemaker nucleus - uEOD unit electric organ discharge     

Immunohistochemical identification of substance P in cutaneous sensory organs (electroreceptors) of gymnotid teleost fish

Summary The distribution of the neuropeptide substance P, which is considered to be a neurotransmitter or neuromodulator of the central nervous system, has been studied in the cutaneous electroreceptor organs (tuberous and ampullary organs) of 3 species of gymnotid fish: Apteronotus leptorhynchus, Eigenmannia virescens and Sternopygus sp. Immunohistochemical data have revealed that substance P is never present in the afferent fibers but is specifically localized in the electroreceptors of the three species examined. Substane P immunoreactivity is strictly localized in the sensory cells of the ampullary organs of all three species and in those of the tuberous organs of Apteronotus leptorhynchus and Sternopygus sp. In contrast, weak substance P immunoreactivity was observed only in certain tuberous sensory cells of Eigenmannia. Substance P immunoreactivity was also found in the accessory cells of certain organs: it was detected in the two types of accessory cells of the tuberous organs of Eigemmannia virescens, in the accessory cells type 2 of the tuberous organs of Sternopygus sp., and in all accessory cells of ampullary organs of Sternopygus sp. and Apteronotus leptorhynchus. In Sternopygus sp., positive staining was only evident if the substance P antibody was used at low concentration. Immunoreactivity for substance P in the sensory cells suggests that it has a transmitter or modulator function in these electroreceptors; the presence of substance P in the accessory cells remains to be explained.     

Spinal mechanisms of electric organ discharge synchronization in Gymnotus carapo

Summary The duration of the electric organ discharge (EOD) in Gymnotus carapo is brief and independent of fish size. Spinal mechanisms involved in electrocyte synchronization were explored by recording spontaneous action potentials of single fibers from the electromotor bulbospinal tract (EBST). Using the field potential of the medullary electromotor nucleus (MEN) as a temporal reference we calculated the orthodromic conduction velocity (CV) of these fibers (range: 10.7–91 m/s).The CVs (in m/s) of fibers recorded at the same level of the spinal cord were significantly different in small and large fish; this difference disappeared when CV were expressed as percentage of body length/ms. Plotting these values against conduction distance (also in %) showed that low CV fibers predominate in the rostral cord while only fast fibers are found at distal levels. Moreover, antidromic stimulation of the distal cord was only effective on high CV fibers. The orthodromic CVs in the distal portion of the recorded fibers were calculated by collision experiments; no significant differences were found between proximal and distal portions.The spatial distribution of CV values within the EBST is proposed to play the main role in synchronizing the electromotoneurons' activity along the spinal cord.Abbreviations EOD electric organ discharge - EO electric organ - EBST electromotor bulbospinal tract - MEN medullary electromotor nucleus - CV conduction velocity - EMN electromotoneuron     

Electric organ activation in Gymnotus carapo: Spinal origin and peripheral mechanisms

A new technique of multiple-air-gap recording was developed to study the EO activation process in Gymnotus carapo. Using this technique, the spatiotemporal pattern of electromotive force generation was investigated in normal and spinal-lesioned animals.Our data indicate that the EOD may be considered as the result of the sequential activation of 3 defined portions of the EO: the abdominal portion (included in the rostral 25% of the fish body), the central portion (comprising the intermediate 50% of the fish body) and the tail portion (the caudal 25% of the fish body). The EOD generated at each portion is characterized by: 1) timing respect to the pacemaker nucleus discharge, 2) speed of progression within the region, 3) waveform, and 4) magnitude.Spinal sections demonstrated that EMNs serving relatively small portions of the EO are widely distributed (convergence) and that surgical exclusion of relatively small portions of the spinal cord diminishes the amplitude of the EOD along an extended portion of the EO (divergence).Abbreviations EMF electromotive force - EMN electromotor-neurons - EO electric organ - EOD electric organ discharge - PMNFP pacemaker nucleus field potential - PEN posterior electromotor nerve - PNA peripheral neural activity     

Social electric signals in freely moving dyads of <Emphasis Type="Italic">Brachyhypopomus pinnicaudatus</Emphasis>

Brachyhypopomus pinnicaudatus (pulse-type weakly electric fish) is a gregarious species that displays reproductive behavior and agonistic encounters between males only during the breeding season. During social interactions, in addition to its basal electric organ discharge (EOD), fish emit social electric signals (SESs) in the contexts of reproduction and intrasexual aggression. We reproduced natural behavior in laboratory settings: SESs recorded in the field are indistinguishable from those observed in our experimental setup. SESs are nocturnal, change seasonally and exhibit sexual dimorphism. This study provides an exhaustive characterization and classification of SESs produced by males and females during the breeding season. In male–female dyads, males produce accelerations and chirps while females interrupt their EODs. The same SESs are observed in male–male dyads. We present a novel, thorough classification of male chirps into four independent types (A, B, C, and M) based on their duration and internal structure. The type M chirp is only observed in male–male dyads. Chirps and interruptions, both in male–female and male–male dyads, are emitted in bouts, which are also grouped throughout the night. Our data suggest the existence of a sophisticated electric dialog during reproductive and aggressive interaction whose precise timing and behavioral significance are being investigated.     

Waveform generation in Rhamphichthys rostratus (L.) (Teleostei,Gymnotiformes)

Rhamphichthys rostratus (L.) emits brief pulses (2 ms) repeated very regularly at 50 Hz. The electric organ shows a heterogeneous distribution of the electrocyte tubes and the occurrence of three electrocyte types (caudally innervated, rostrally innervated and marginallycaudally innervated). In the sub-opercular region the electric organ consists of a pair of tubes containing only caudally innervated electrocytes. At the abdominal region the EO consists of three pairs of tubes. Each pair contains one of the described electrocyte types. The number of electrocyte tubes increases toward the tail to reach nine or ten pairs in the most caudal segments. In the intermediate region most tubes contain doubly innervated electrocytes except the ventral pair that contains caudally innervated electrocytes. The caudal 25% contains exclusively caudally innervated electrocytes. The electric organ discharge consists of five wave components (V1 to V5). Electrophysiological data are consistent with the hypothesis that V1 results from the activity of the rostral faces of rostrally innervated electrocytes. V2 results from the activities of rostral faces of marginally-caudally innervated electrocytes while V3 results from the activities of caudal faces of most electrocytes. Curarization experiments demonstrated that V4 and V5 result from action potential invasion and are not directly elicited by neural activity.Abbreviations AEN1 anterior electromotor nerve 1 - AEN2 anterior electromotor nerve 2 - BMB boraxic methylene blue - CIE caudally innervated electrocytes - EMF electromotive force - EO electric organ - EOD electric organ discharge - I current amplitude - MCIE marginally-caudally innervated electrocytes - MT medial tubes - PEN posterior electromotor nerve - R _n internal impedance - RIE rostrally innervated electrocytes - Rl load resistor - SAT short abdominal tubes - V voltage amplitude     

Influence of temperature and reproductive state upon the jamming avoidance response in the pulse-type electric fish <Emphasis Type="Italic">Brachyhypopomus pinnicaudatus</Emphasis>

The electric organ discharge (EOD) in Brachyhypopomus pinnicaudatus is modified by temperature and reproductive state. We studied the influence of these variables upon a complex behavior, the jamming avoidance response (JAR). Experiments were performed in non-reproductive fish and in two groups of fish after the induction of reproductive state (by nature or by acclimation at 28°C). JARs were elicited at 20 and 30°C by free-run electric stimuli with different Ls (interval difference between stimulus and EOD). In non-reproductive fish, JARs induced by stimuli with +Ls showed temperature sensitivity, with smaller responses at 30°C. Conversely, similar JARs were obtained at both temperatures in reproductive fish. These observations were replicated in curarized preparations. Stimuli with –Ls were almost ineffective in non-reproductive fish at 30°C, whereas adequate JARs were shown by reproductive fish. Phase-locked stimuli were used to evaluate the duration of the low-threshold electrosensory periods preceding and following the EOD. In non-reproductive fish, the temperature step induced a shortening of these periods. The opposite effect was observed in reproductive fish, probably explaining the differences in JAR capability. A prolongation of the low-threshold periods would favor the perception of electrocommunication signals during courtship. JAR changes would be a consequence of this adaptation.