SUMMARY Electric organs (EOs) have evolved independently in vertebrates six times from skeletal muscle (SM). The transcriptional changes accompanying this developmental transformation are not presently well understood. Mormyrids and gymnotiforms are two highly convergent groups of weakly electric fish that have independently evolved EOs: while much is known about development and gene expression in gymnotiforms, very little is known about development and gene expression in mormyrids. This lack of data limits prospects for comparative work. We report here on the characterization of 28 differentially expressed genes between SM and EO tissues in the mormyrid Brienomyrus brachyistius , which were identified using suppressive subtractive hybridization (SSH). Forward and reverse SSH was performed on tissue samples of EO and SM resulting in one cDNA library enriched with mRNAs expressed in EO, and a second library representing mRNAs unique to SM. Nineteen expressed sequence tags (ESTs) were identified in EO and nine were identified in SM using BLAST searching of Danio rerio sequences available in NCBI databases. We confirmed differential expression of all 28 ESTs using RT-PCR. In EO, these ESTs represent four classes of proteins: (1) ion pumps, including the α- and β-subunits of Na + /K + -ATPase, and a plasma membrane Ca 2+ -ATPase; (2) Ca 2+ -binding protein S100, several parvalbumin paralogs, calcyclin-binding protein and neurogranin; (3) sarcomeric proteins troponin I, myosin heavy chain and actin-related protein complex subunit 3 (Arcp3); and (4) the transcription factors enhancer of rudimentary homolog (ERH) and myocyte enhancer factor 2A (MEF2A). Immunohistochemistry and western blotting were used to demonstrate the translation of seven proteins (myosin heavy chain, Na + /K + -ATPase, plasma membrane Ca 2+ -ATPase, MEF2, troponin and parvalbumin) and their cellular localization in EO and SM. Our findings suggest that mormyrids express several paralogs of muscle-specific genes and the proteins they encode in EOs, unlike gymnotiforms, which may post-transcriptionally repress several sarcomeric proteins. In spite of the similarity in the physiology and function of EOs in mormyrids and gymnotiforms, this study indicates that the mechanisms of development in the two groups may be considerably different.

SUMMARY Mormyrid electric fish rely on the waveform of their electric organ discharges (EODs) for communicating species, sex, and social status, while they use the sequences of pulse intervals (SPIs) for communicating rapidly changing behavioral states and motivation. Little is known of electric signaling during courtship behavior because of two major difficulties: (1) the fish are not easily bred in captivity and (2) there is no reliable means of separating electric signals from several individuals in natural communication settings. Through simulating artificial rain conditions, we have successfully induced courtship and succeeded in breeding a mormyrid electric fish( Brienomyrus brachyistius ) in the laboratory. We have also developed a system of video recording and editing combined with cross correlation analysis to precisely record and view behavior and separate EODs from two individuals in non-breeding and breeding contexts. Knowing the electrical and motor patterns during courtship allows for further exploration of topics such as mate choice and neural basis of pattern generation in these fish. Here we describe nine common motor displays and 11 SPIs. Analysis of frequency of occurrences suggests that some SPI patterns are sex and season specific. We also observed electrical duetting called `rasp matching' during courtship signaling among pairs; males and females exchange `rasps' and`bursts', respectively, in alternation. Our study employs new techniques to separate and document SPIs in the context of courtship. We show that some SPIs correlate with specific behavioral acts around the time of spawning.

SUMMARY Polymorphism in an animal communication channel provides a framework for studying proximate rules of signal design as well as ultimate mechanisms of signal diversification. Reproductively isolated mormyrid fishes from Gabon's Brienomyrus species flock emit distinctive electric organ discharges(EODs) thought to function in species and sex recognition. Species boundaries and EODs appear congruent in these fishes, with the notable exception of three morphs designated types I, II and III. Within the species flock, these morphs compose a monophyletic group that has recently been called the magnostipes complex. Co-occurring morphs of this complex express distinctive EODs, yet they appear genetically indistinguishable at several nuclear loci. In this study, we investigated EOD discrimination by these morphs using both behavioral and physiological experiments. During the breeding season, wild-caught type I and type II males showed evidence that they can discriminate their own morph's EOD waveform from that of a sympatric and genetically distinct reference species. However, we found that type I and type II males exhibited an asymmetry in unconditioned responses to paired playback of EODs recorded from type I versus type II females. Males of the type II morph responded preferentially to EODs of type II females,whereas type I males did not appear to discriminate homotypic and heterotypic EODs in our experimental paradigm. Part of this behavioral asymmetry may have resulted from a previously undetected difference in adult size, which may have enhanced apparent discrimination by the smaller morph (type II) due to a relatively higher risk of injury from the larger morph (type I). Knollenorgan receptors, which mediate electrical communication in mormyrids, showed similar spectral tuning in type I and type II. These electroreceptors coded temporal features of any single magnostipes -complex EOD with similar patterns of time-locked spikes in both morphs. By contrast, Knollenorgans exhibited distinctive responses to different EOD waveforms. These results suggest that discrete EOD variation in this rapidly diversifying complex is functional in terms of morph-specific advertisement and recognition. Time-domain signal divergence has outpaced frequency-domain divergence between sympatric morphs,requiring little to no change in receptor response properties. We discuss our findings in light of a model for EOD time-coding by the Knollenorgan pathway,as well as evolutionary hypotheses concerning sympatric signal diversification in the magnostipes complex.

SUMMARY Like all mormyrid fish, Brienomyrus brachyistius produces an electric organ discharge (EOD) with a constant waveform and variable sequence of pulse intervals (SPI). Periodic bursts fall into two display categories termed `scallops' and `accelerations', with a third category termed `rasps'that appears to combine the two. The medullary EOD command nucleus (CN)receives excitatory input from the midbrain precommand nucleus (PCN) and the thalamic dorsal posterior nucleus (DP), both of which are regulated by a recurrent inhibitory projection from the ventroposterior nucleus of the torus semicircularis (VP). We tested the following hypotheses: (1) PCN and DP are responsible for generating different burst types (scallops and accelerations,respectively), (2) differences in the strength of recurrent inhibition are related to physiological differences between PCN and DP and (3) recurrent inhibition regulates the resting electromotor rhythm, while disinhibition releases PCN and DP, allowing them to generate bursts. Iontophoresis of the excitatory neurotransmitter l -glutamate ( l -Glu) into DP led to acceleration-like output patterns, while in PCN it led to scallop-like output patterns. Iontophoresis of the inhibitory neurotransmitterγ-amino-butyric acid (GABA) into DP and PCN led to an elongation of intervals, as did iontophoresis of l -Glu into VP. Iontophoresis of the GABA A receptor blocker bicuculline methiodide (BMI) into DP and PCN induced repetitive bursting behavior and eliminated differences in the effects of l -Glu iontophoresis in the two nuclei. These results support our three hypotheses, suggesting that production of different communication behaviors may be regulated by spatially distinct groups of neurons, and recurrent inhibition and disinhibition may play an active role in driving and shaping such behaviors.